Difference between revisions of "Team:TJUSLS China/Description"
| Line 7: | Line 7: | ||
<meta http-equiv="X-UA-Compatible" content="ie=edge"> | <meta http-equiv="X-UA-Compatible" content="ie=edge"> | ||
<link rel="stylesheet" type="text/css" | <link rel="stylesheet" type="text/css" | ||
| − | + | href="https://2019.igem.org/Team:TJUSLS_China/assets/vendors/bootstrap/gridcss?action=raw&ctype=text/css"> | |
<link rel="stylesheet" type="text/css" | <link rel="stylesheet" type="text/css" | ||
| − | + | href="https://2019.igem.org/Team:TJUSLS_China/assets/vendors/magnific-popup/magnific-popupmincss?action=raw&ctype=text/css"> | |
<link rel="stylesheet" type="text/css" | <link rel="stylesheet" type="text/css" | ||
| − | + | href="https://2019.igem.org/Team:TJUSLS_China/assets/vendors/swiper/swipercss?action=raw&ctype=text/css"> | |
<link rel="stylesheet" type="text/css" | <link rel="stylesheet" type="text/css" | ||
| − | + | href="https://2019.igem.org/Team:TJUSLS_China/assets/vendors/wow/animatecss?action=raw&ctype=text/css"> | |
<!-- App & fonts--> | <!-- App & fonts--> | ||
<link rel="stylesheet" type="text/css" id="app-stylesheet" | <link rel="stylesheet" type="text/css" id="app-stylesheet" | ||
| − | + | href="https://2019.igem.org/Team:TJUSLS_China/assets/css/main?action=raw&ctype=text/css"> | |
<script type="text/javascript" | <script type="text/javascript" | ||
| − | + | src="https://2019.igem.org/Team:TJUSLS_China/assets/js/srcjs?action=raw&ctype=text/javascript"></script> | |
<script type="text/javascript" | <script type="text/javascript" | ||
| − | + | src="https://2019.igem.org/Team:TJUSLS_China/assets/js/jqueryminjs?action=raw&ctype=text/javascript"></script> | |
<script type="text/javascript" | <script type="text/javascript" | ||
| − | + | src="https://2019.igem.org/Team:TJUSLS_China/js?action=raw&ctype=text/javascript"></script> | |
<script type="text/javascript" | <script type="text/javascript" | ||
| − | + | src="https://2019.igem.org/Team:TJUSLS_China/assets/js/chartjs?action=raw&ctype=text/javascript"></script> | |
<title>Description</title> | <title>Description</title> | ||
<style> | <style> | ||
| Line 91: | Line 91: | ||
} | } | ||
| − | #title>h1, | + | #title > h1, |
| − | #title-cover>h1 { | + | #title-cover > h1 { |
font-size: 130px; | font-size: 130px; | ||
font-weight: bolder; | font-weight: bolder; | ||
| Line 186: | Line 186: | ||
/* .menuHolder ul li { | /* .menuHolder ul li { | ||
| − | + | border-radius: 0 0 300px 0; | |
| − | + | width: 0; | |
| − | + | height: 0; | |
| − | + | } */ | |
.menuHolder ul li a { | .menuHolder ul li a { | ||
| Line 215: | Line 215: | ||
/*画第一个圆圈*/ | /*画第一个圆圈*/ | ||
| − | .menuHolder li.s1>a { | + | .menuHolder li.s1 > a { |
position: absolute; | position: absolute; | ||
display: block; | display: block; | ||
| Line 225: | Line 225: | ||
/*画第二个圆圈*/ | /*画第二个圆圈*/ | ||
| − | .menuHolder li.s2>a { | + | .menuHolder li.s2 > a { |
position: absolute; | position: absolute; | ||
display: block; | display: block; | ||
| Line 254: | Line 254: | ||
/*绘制第二层的菜单*/ | /*绘制第二层的菜单*/ | ||
| − | .menuHolder li.s2:nth-of-type(8)>a { | + | .menuHolder li.s2:nth-of-type(8) > a { |
background: #888; | background: #888; | ||
transform: rotate(78.25deg); | transform: rotate(78.25deg); | ||
} | } | ||
| − | .menuHolder li.s2:nth-of-type(7)>a { | + | .menuHolder li.s2:nth-of-type(7) > a { |
background: #999; | background: #999; | ||
transform: rotate(67.5deg); | transform: rotate(67.5deg); | ||
} | } | ||
| − | .menuHolder li.s2:nth-of-type(6)>a { | + | .menuHolder li.s2:nth-of-type(6) > a { |
background: #aaa; | background: #aaa; | ||
transform: rotate(56.25deg); | transform: rotate(56.25deg); | ||
} | } | ||
| − | .menuHolder li.s2:nth-of-type(5)>a { | + | .menuHolder li.s2:nth-of-type(5) > a { |
background: #bbb; | background: #bbb; | ||
transform: rotate(45deg); | transform: rotate(45deg); | ||
} | } | ||
| − | .menuHolder li.s2:nth-of-type(4)>a { | + | .menuHolder li.s2:nth-of-type(4) > a { |
background: #ccc; | background: #ccc; | ||
transform: rotate(33.75deg); | transform: rotate(33.75deg); | ||
} | } | ||
| − | .menuHolder li.s2:nth-of-type(3)>a { | + | .menuHolder li.s2:nth-of-type(3) > a { |
background: #ddd; | background: #ddd; | ||
transform: rotate(22.5deg); | transform: rotate(22.5deg); | ||
} | } | ||
| − | .menuHolder li.s2:nth-of-type(2)>a { | + | .menuHolder li.s2:nth-of-type(2) > a { |
background: #eee; | background: #eee; | ||
transform: rotate(11.25deg); | transform: rotate(11.25deg); | ||
} | } | ||
| − | .menuHolder li.s2:nth-of-type(1)>a { | + | .menuHolder li.s2:nth-of-type(1) > a { |
background: #fff; | background: #fff; | ||
transform: rotate(0deg); | transform: rotate(0deg); | ||
| Line 295: | Line 295: | ||
/*绘制Parts的子菜单*/ | /*绘制Parts的子菜单*/ | ||
| − | .menuHolder .a6 li:nth-of-type(4)>a { | + | .menuHolder .a6 li:nth-of-type(4) > a { |
background: #999; | background: #999; | ||
transform: rotate(67.5deg); | transform: rotate(67.5deg); | ||
} | } | ||
| − | .menuHolder .a6 li:nth-of-type(3)>a { | + | .menuHolder .a6 li:nth-of-type(3) > a { |
background: #bbb; | background: #bbb; | ||
transform: rotate(45deg); | transform: rotate(45deg); | ||
} | } | ||
| − | .menuHolder .a6 li:nth-of-type(2)>a { | + | .menuHolder .a6 li:nth-of-type(2) > a { |
background: #ddd; | background: #ddd; | ||
transform: rotate(22.5deg); | transform: rotate(22.5deg); | ||
} | } | ||
| − | .menuHolder .a6 li:nth-of-type(1)>a { | + | .menuHolder .a6 li:nth-of-type(1) > a { |
background: #fff; | background: #fff; | ||
transform: rotate(0deg); | transform: rotate(0deg); | ||
| Line 317: | Line 317: | ||
/*绘制Team的子菜单*/ | /*绘制Team的子菜单*/ | ||
| − | .menuHolder .a5 li:nth-of-type(3)>a { | + | .menuHolder .a5 li:nth-of-type(3) > a { |
background: #aaa; | background: #aaa; | ||
transform: rotate(60deg); | transform: rotate(60deg); | ||
} | } | ||
| − | .menuHolder .a5 li:nth-of-type(2)>a { | + | .menuHolder .a5 li:nth-of-type(2) > a { |
background: #ccc; | background: #ccc; | ||
transform: rotate(30deg); | transform: rotate(30deg); | ||
} | } | ||
| − | .menuHolder .a5 li:nth-of-type(1)>a { | + | .menuHolder .a5 li:nth-of-type(1) > a { |
background: #fff; | background: #fff; | ||
transform: rotate(0deg); | transform: rotate(0deg); | ||
| Line 333: | Line 333: | ||
/*绘制Project的子菜单*/ | /*绘制Project的子菜单*/ | ||
| − | .menuHolder .a3 li:nth-of-type(1)>a { | + | .menuHolder .a3 li:nth-of-type(1) > a { |
background: #fff; | background: #fff; | ||
transform: rotate(0deg); | transform: rotate(0deg); | ||
} | } | ||
| − | .menuHolder .a3 li:nth-of-type(8)>a { | + | .menuHolder .a3 li:nth-of-type(8) > a { |
background: #888; | background: #888; | ||
transform: rotate(78.75deg); | transform: rotate(78.75deg); | ||
| Line 344: | Line 344: | ||
} | } | ||
| − | .menuHolder .a3 li:nth-of-type(7)>a { | + | .menuHolder .a3 li:nth-of-type(7) > a { |
background: #999; | background: #999; | ||
transform: rotate(67.5deg); | transform: rotate(67.5deg); | ||
} | } | ||
| − | .menuHolder .a3 li:nth-of-type(6)>a { | + | .menuHolder .a3 li:nth-of-type(6) > a { |
background: #aaa; | background: #aaa; | ||
transform: rotate(56.25deg); | transform: rotate(56.25deg); | ||
} | } | ||
| − | .menuHolder .a3 li:nth-of-type(5)>a { | + | .menuHolder .a3 li:nth-of-type(5) > a { |
background: #bbb; | background: #bbb; | ||
transform: rotate(45deg); | transform: rotate(45deg); | ||
} | } | ||
| − | .menuHolder .a3 li:nth-of-type(4)>a { | + | .menuHolder .a3 li:nth-of-type(4) > a { |
background: #ccc; | background: #ccc; | ||
transform: rotate(33.75deg); | transform: rotate(33.75deg); | ||
} | } | ||
| − | .menuHolder .a3 li:nth-of-type(3)>a { | + | .menuHolder .a3 li:nth-of-type(3) > a { |
background: #ddd; | background: #ddd; | ||
transform: rotate(22.5deg); | transform: rotate(22.5deg); | ||
} | } | ||
| − | .menuHolder .a3 li:nth-of-type(2)>a { | + | .menuHolder .a3 li:nth-of-type(2) > a { |
background: #eee; | background: #eee; | ||
transform: rotate(11.25deg); | transform: rotate(11.25deg); | ||
} | } | ||
| − | .menuHolder .a7 li:nth-of-type(2)>a { | + | .menuHolder .a7 li:nth-of-type(2) > a { |
background: #aaa; | background: #aaa; | ||
transform: rotate(45deg); | transform: rotate(45deg); | ||
} | } | ||
| − | .menuHolder .a7 li:nth-of-type(1)>a { | + | .menuHolder .a7 li:nth-of-type(1) > a { |
background: #fff; | background: #fff; | ||
transform: rotate(0deg); | transform: rotate(0deg); | ||
| Line 395: | Line 395: | ||
/*鼠标滑过改变背景颜色和文字颜色*/ | /*鼠标滑过改变背景颜色和文字颜色*/ | ||
| − | .menuHolder ul li:hover>a { | + | .menuHolder ul li:hover > a { |
background: #11253b; | background: #11253b; | ||
color: #fff; | color: #fff; | ||
} | } | ||
| − | .menuHolder li.s2:hover>a { | + | .menuHolder li.s2:hover > a { |
background: #11253b; | background: #11253b; | ||
color: #fff; | color: #fff; | ||
} | } | ||
| − | .menuHolder .a6 li:hover>a { | + | .menuHolder .a6 li:hover > a { |
background: #11253b; | background: #11253b; | ||
color: #fff; | color: #fff; | ||
} | } | ||
| − | .menuHolder .a5 li:hover>a { | + | .menuHolder .a5 li:hover > a { |
background: #11253b; | background: #11253b; | ||
color: #fff; | color: #fff; | ||
} | } | ||
| − | .menuHolder .a3 li:hover>a { | + | .menuHolder .a3 li:hover > a { |
background: #11253b; | background: #11253b; | ||
color: #fff; | color: #fff; | ||
} | } | ||
| − | .menuHolder .a7 li:hover>a { | + | .menuHolder .a7 li:hover > a { |
background: #11253b; | background: #11253b; | ||
color: #fff; | color: #fff; | ||
| Line 546: | Line 546: | ||
} | } | ||
| − | #mw-content-text>p { | + | #mw-content-text > p { |
display: none !important; | display: none !important; | ||
} | } | ||
| Line 844: | Line 844: | ||
<body> | <body> | ||
| − | + | <div id="main" class="fixed"> | |
| − | + | <div id="navbar" style="padding-top: 6px;"> | |
| − | + | <div id="links"> | |
| − | + | <a href="#id-6">Reference</a> | |
| − | + | <a href="#id-5">Aim & Meanings</a> | |
| − | + | <a href="#id-4">Methods</a> | |
| − | + | <a href="#id-3">Inspirations</a> | |
| − | + | <a href="#id-2">Background</a> | |
| − | + | <a href="#id-1">Overview</a> | |
| − | + | </div> | |
| − | + | <div class="menuHolder"> | |
| − | + | <div class="menuWindow"> | |
| − | + | <ul class="p1"> | |
| − | + | <li class="s1"> | |
| − | + | <a href="http://2019.igem.org/Team:TJUSLS_China/Description"> | |
| − | + | <img src="https://2019.igem.org/wiki/images/4/48/T--TJUSLS_China--logo.svg" alt="TJUSLS" | |
| − | + | id="navbar-logo"> | |
| − | + | </a> | |
| − | + | <ul class="p2" style="transform: rotate(0deg);"> | |
| − | + | <li class="s2"><a href="http://2019.igem.org/Team:TJUSLS_China"><span>HOME</span></a> | |
| − | + | </li> | |
| − | + | <li class="s2"><a | |
| − | + | href="http://2019.igem.org/Team:TJUSLS_China/Description"><span>PROJECT</span></a> | |
| − | + | <ul class="p3 a3"> | |
| − | + | <li><a href="http://2019.igem.org/Team:TJUSLS_China/Description">Description</a> | |
| − | + | </li> | |
| − | + | <li><a href="http://2019.igem.org/Team:TJUSLS_China/Design">Design</a></li> | |
| − | + | <li><a href="http://2019.igem.org/Team:TJUSLS_China/Experiments">Experiments</a> | |
| − | + | </li> | |
| − | + | <li><a href="http://2019.igem.org/Team:TJUSLS_China/Notebook">Notebook</a></li> | |
| − | + | <li><a href="http://2019.igem.org/Team:TJUSLS_China/Demonstrate">Demonstrate</a> | |
| − | + | </li> | |
| − | + | <li><a href="http://2019.igem.org/Team:TJUSLS_China/Improve">Improve</a></li> | |
| − | + | <li><a | |
| − | + | href="http://2019.igem.org/Team:TJUSLS_China/Characterization">Characterization</a> | |
| − | + | </li> | |
| − | + | <li><a href="http://2019.igem.org/Team:TJUSLS_China/Results">Results</a></li> | |
| − | + | </ul> | |
| − | + | </li> | |
| − | + | <li class="s2"><a | |
| − | + | href="http://2019.igem.org/Team:TJUSLS_China/Safety"><span>SAFETY</span></a> | |
| − | + | </li> | |
| − | + | <li class="s2"><a | |
| − | + | href="http://2019.igem.org/Team:TJUSLS_China/Parts"><span>PARTS</span></a> | |
| − | + | <ul class="p3 a6"> | |
| − | + | <li><a href="http://2019.igem.org/Team:TJUSLS_China/Parts">Overview</a></li> | |
| − | + | <li><a href="http://2019.igem.org/Team:TJUSLS_China/Basic_Part">Basic parts</a> | |
| − | + | </li> | |
| − | + | <li><a href="http://2019.igem.org/Team:TJUSLS_China/Composite_Part">Composite | |
| − | + | parts</a></li> | |
| − | + | <li><a href="http://2019.igem.org/Team:TJUSLS_China/Part_Collection">Part | |
| − | + | collection</a></li> | |
| − | + | </ul> | |
| − | + | </li> | |
| − | + | <li class="s2"><a | |
| − | + | href="http://2019.igem.org/Team:TJUSLS_China/Model"><span>MODEL</span></a> | |
| − | + | </li> | |
| − | + | <li class="s2"><a | |
| − | + | href="http://2019.igem.org/Team:TJUSLS_China/Human_Practices"><span>HP</span></a> | |
| − | + | <ul class="p3 a7"> | |
| − | + | <li><a href="http://2019.igem.org/Team:TJUSLS_China/Human_Practices">Human | |
| − | + | practices</a></li> | |
| − | + | <li><a href="http://2019.igem.org/Team:TJUSLS_China/Public_Engagement">Education | |
| − | + | and engagement</a></li> | |
| − | + | </ul> | |
| − | + | </li> | |
| − | + | <li class="s2"><a href="http://2019.igem.org/Team:TJUSLS_China/Medal"><span>JUDGING | |
FORM</span></a> | FORM</span></a> | ||
| − | + | <ul class="p3 a5"> | |
| − | + | <li><a href="http://2019.igem.org/Team:TJUSLS_China/Medal">Medal | |
| − | + | requirements</a> | |
| − | + | </li> | |
| − | + | <li><a href="http://2019.igem.org/Team:TJUSLS_China/Measurement">Measurement</a> | |
| − | + | </li> | |
| − | + | <li><a href="http://igem.org/2019_Judging_Form?team=TJUSLS_China">Judging | |
| − | + | Form</a> | |
| − | + | </li> | |
| − | + | </ul> | |
| − | + | </li> | |
| − | + | <li class="s2 b6"><a | |
| − | + | href="http://2019.igem.org/Team:TJUSLS_China/Team"><span>TEAM</span></a> | |
| − | + | <ul class="p3 a5"> | |
| − | + | <li><a href="http://2019.igem.org/Team:TJUSLS_China/Team">Team members</a></li> | |
| − | + | <li> | |
| − | + | <a | |
href="http://2019.igem.org/Team:TJUSLS_China/Collaborations">Collaborations</a> | href="http://2019.igem.org/Team:TJUSLS_China/Collaborations">Collaborations</a> | ||
| − | + | </li> | |
| − | + | <li><a | |
| − | + | href="http://2019.igem.org/Team:TJUSLS_China/Attributions">Attributions</a> | |
| − | + | </li> | |
| − | + | </ul> | |
| − | + | </li> | |
| − | + | </ul> | |
| − | + | </li> | |
| − | + | </ul> | |
| − | + | ||
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div id="outerdiv" | |
| − | + | style="text-align: center;position: fixed;z-index: 1000;top: 0;left: 0;width: 100%;height: 100%;background-color: rgba(255,255,255,.9);"> | |
| − | + | <div style="padding-top: 5%;font-size: 20px;color: #11253b;font-weight: 700">click anywhere to close</div> | |
| − | + | <img id="bigimg" | |
| − | + | style="height: auto;width: 46.6325%;border: 0;margin: auto;position: absolute;top: 0;bottom: 0;left: 0;right: 0;" | |
| − | + | src=""/> | |
| − | + | </div> | |
| − | + | <script> | |
| − | + | //图片放大 | |
| − | + | $("#outerdiv").hide(); | |
| − | + | $(function () { | |
| − | + | $("img").each(function () { | |
| − | + | var id = $(this).attr("id"); | |
| − | + | if (id != "navbar-logo") { | |
| − | + | $(this).mouseover(function () { | |
| − | + | $(this).css("cursor", "pointer"); | |
| + | }); | ||
| − | + | $(this).click(function () { | |
| − | + | var _this = $(this);//将当前的pimg元素作为_this传入函数 | |
| − | + | imgShow("#outerdiv", "#bigimg", _this); | |
| − | + | }); | |
| − | + | } | |
| − | } | + | |
}); | }); | ||
| + | }); | ||
| − | + | function imgShow(outerdiv, bigimg, _this) { | |
| − | + | var src = _this.attr("src");//获取当前点击的pimg元素中的src属性 | |
| − | + | $("#outerdiv").attr("display", "block"); | |
| − | + | $("#bigimg").attr("src", src);//设置#bigimg元素的src属性 | |
| − | + | $("#outerdiv").fadeIn("fast"); | |
| − | + | $("#outerdiv").click(function () {//再次点击淡出消失弹出层 | |
| − | + | $(this).fadeOut("fast"); | |
| − | + | }); | |
| − | + | } | |
| − | + | </script> | |
| − | + | <div style="padding-top: 16px"> | |
| − | + | <section class="py-7 py-md-0 bg-hero" | |
| − | + | style="background-image: url(https://2019.igem.org/wiki/images/4/4a/T--TJUSLS_China--description-headpic.jpg)" | |
| − | + | id="home"> | |
| − | + | <div class="container1"> | |
| − | + | <div class="row1 vh-md-100"> | |
| − | + | <div class="col-md-7 mx-auto my-auto text-center" style="text-align: center"> | |
| − | + | <h1 | |
style="color: white;border:none;font-size: 7.75rem;font-weight: 700;height:100%;padding-top: 30%"> | style="color: white;border:none;font-size: 7.75rem;font-weight: 700;height:100%;padding-top: 30%"> | ||
| − | + | Description</h1> | |
| − | + | ||
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="cover-arrow bounce d-none d-md-block"> | |
| − | + | <a href="#id-1" class="page-scroll"><em class="text-white" data-feather="chevrons-down"></em></a> | |
| − | + | </div> | |
| − | + | </section> | |
| − | + | </div> | |
| − | + | <section class="awe-section" style="background: white" id="id-1"> | |
| − | + | <div class="container"> | |
| − | + | <div class="row"> | |
| − | + | <!-- title --> | |
| − | + | <div class="title"> | |
| − | + | <h2 class="title__title" style="color: #11253b">Overview</h2> | |
| − | + | </div><!-- End / title --> | |
| − | + | <!-- about --> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10 "> | |
| − | + | <div class="about__words"> | |
| − | + | <p style="color: black; font-size:18px;line-height: 2.1"> | |
| − | + | We focus our project on hitting Achilles’ Heel of metallo-beta-lactamases (MBLs), a | |
| − | + | vital class of beta-lactamases without available clinical inhibitors, produced by | |
| − | + | drug-resistant pathogens. We want to obtain the broad-spectrum inhibitor of MBLs and | |
| − | + | lay the foundation for clinical trials in the future. | |
| − | + | </p> | |
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| + | <div class="about"> | ||
| − | + | <div class="col-lg-12 "> | |
| − | + | <div style="text-align: center" class="about__img"> | |
| − | + | <img style="width: 70%;height: 70%;" | |
| − | + | src="https://2019.igem.org/wiki/images/f/fa/T--TJUSLS_China--description-0.png"> | |
| − | + | </div> <!-- 1260 * 750 --> | |
| − | + | ||
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row" style="padding-top: 5%"> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1"></div> | |
| − | + | <div class="col-lg-10"> | |
| − | + | <div class="about__words"> | |
| − | + | <p style="color: black; font-size:18px;line-height: 2.1"> | |
| − | + | Firstly we used synthetic biology methods to express a series of MBLs in E. coli. | |
| − | + | Besides, a virtual screening model has been built to discover the therapeutically | |
| − | + | effective compounds, in order to predict potential inhibitors. Next, effective compounds | |
| − | + | were screened via high-throughput screening with fluorescent probe (CDC-1) from FDA | |
| − | + | approved drug libraries and traditional Chinese medicine libraries. Also we assessed | |
| − | + | their inhibitory ability in living bacterial cells by UV-vis spectroscopy. Finally we | |
| − | + | have obtained a series of effective inhibitors of MBLs, including an excellent | |
| − | + | broad-spectrum inhibitor, which can prevent living resistant bacteria from hydrolyzing | |
| − | + | beta-lactam antibiotics. We hope these inhibitors can be ideal candidates for | |
| − | + | therapeutics for diseases caused by drug-resistant pathogens. | |
| + | </p> | ||
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
| − | </section> | + | </div> |
| − | + | </section> | |
| − | + | <section class="awe-section" style="background: white;padding-top: 0px" id="id-2"> | |
| − | + | <div class="container"> | |
| − | + | <div class="row"> | |
| − | + | <div class="about"> | |
| − | + | <div class="title"> | |
| − | + | <h2 class="title__title" style="color: #11253b">Background</h2> | |
| − | + | </div><!-- End / title --> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10 "> | |
| − | + | <div class="about__words"> | |
| − | + | <h2 class="fw-600 fz-30" style="color: #11253b">Metallo-beta-lactamases result in | |
| − | + | beta-lactam resistance</h2> | |
| − | + | <p style="color: black; font-size:18px;line-height: 2.1"> | |
| − | + | The introduction of antibiotics represented one of the most important medical | |
| − | + | interventions in the history of global health, resulting in a dramatic reduction in | |
| − | + | human morbidity and mortality caused by bacterial infections<sup>[1]</sup>. However, | |
| − | + | the overuse of antibiotics has made the production of drug-resistant bacteria a | |
| − | + | global burden. The World Health Organization has named antibiotic resistance as one | |
| − | + | of the three most important public health threats of the 21st century<sup>[2]</sup>. | |
| − | + | </p> | |
| − | + | <p style="color: black; font-size:18px;line-height: 2.1"> | |
| − | + | The beta-lactams are the most successful class of antibiotic drugs but they are | |
| − | + | vulnerable to inactivation by a growing cadre of beta-lactamases that now number | |
| − | + | more than a thousand variants<sup>[3]</sup>. These enzymes cleave the amide bond of | |
| − | + | the beta-lactam ring thus inactivating the antibiotic. | |
| − | + | </p> | |
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10"> | |
| − | + | <div class="about__img" style="text-align: center"> | |
| − | + | <img src="https://2019.igem.org/wiki/images/c/ca/T--TJUSLS_China--description-1.png"> | |
| − | + | <p style="color: black;font-size: 16px">Figure1: The hydrolysis of penicillin.</p> | |
| − | + | </div> <!-- 1260 * 750 --> | |
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1"></div> | |
| − | + | <div class="col-lg-10"> | |
| − | + | <div class="about__words"> | |
| − | + | <p style="color: black; font-size:18px;line-height: 2.1"> | |
| − | + | The Ambler system divides beta-lactamases into four classes, termed A, B, C and D. | |
| − | + | This year, we focus on class B that called metallo-beta-lactamases, or MBLs. Much | |
| − | + | different from other beta-lactamases, MBLs require zinc for activity and catalysis | |
| − | + | does not proceed via a covalent intermediate but rather through direct attack of a | |
| − | + | hydroxide ion that is stabilized by the Zinc ions in the active site<sup>[4]</sup>. | |
| − | + | MBLs confer resistance to most beta-lactam antibiotics including carbapenems(often | |
| − | + | seen as last line drugs)<sup>[3]</sup>, making it a global threat to public health. | |
| − | + | </p> | |
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10"> | |
| − | + | <div class="about__img" style="text-align: center"> | |
| − | + | <img src="https://2019.igem.org/wiki/images/3/3f/T--TJUSLS_China--description-2.png"> | |
| − | + | <p style="color: black;font-size: 16px">Figure2: The active site of blaNDM-1(a typical | |
| − | + | kind of MBLs).</p> | |
| − | + | </div> <!-- 1260 * 750 --> | |
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row" style="padding-top: 6.5%"> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1"></div> | |
| − | + | <div class="col-lg-10 "> | |
| − | + | <h2 class="fw-600 fz-30" style="color: #11253b">Major limitation on relevant therapy：lack of clinical MBLs inhibitors</h2> | |
| − | + | <div class="about__words"> | |
| − | + | <p style="color: black; font-size:18px;line-height: 2.1"> | |
| − | + | At present, due to the huge cost and low return of developing new antibiotics, the | |
| − | + | trend of developing new antibiotics has gradually declined in recent years. | |
| − | + | Alongside improvements to antibiotics themselves, combinations of existing | |
| − | + | beta-lactams with MBLs inhibitors represent the more feasible and common strategy to | |
| − | + | extend the usefulness of these existing beta-lactams.</p> | |
| − | + | ||
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10"> | |
| − | + | <div class="about__img" style="text-align: center"> | |
| − | + | <img src="https://2019.igem.org/wiki/images/f/f7/T--TJUSLS_China--description-new1.png"> | |
| − | + | <p style="color: black;font-size: 16px">Figure3：Schematic diagram of combining | |
| − | + | antibiotic and inhibitor</p> | |
| − | + | </div> <!-- 1260 * 750 --> | |
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row" style="padding-top: 6.5%"> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1"></div> | |
| − | + | <div class="col-lg-10 "> | |
| − | + | <div class="about__words"> | |
| − | + | <p style="color: black; font-size:18px;line-height: 2.1"> | |
| − | + | For other beta-lactamases, a number of these beta-lactam-beta-lactamases inhibitor | |
| − | + | combinations are approved for clinical use previously (for example, | |
| − | + | Amoxicillin-clavulanate is effective in the treatment of community-acquired | |
| − | + | respiratory infections<sup>[5]</sup> )，and many others are in clinical trials. | |
| − | + | </p> | |
| − | + | <p style="color: black; font-size:18px;line-height: 2.1"> | |
| − | + | Unfortunately, the discovery of a clinically useful, specific inhibitor of | |
| − | + | metallo-beta-lactamases is made difficult by the fact that this compound must remain | |
| − | + | inactive towards the human proteins which are members of the metallo-beta-lactamase | |
| − | + | superfamily, or other metallo-enzymes such as the angiotensin converting enzyme. | |
| − | + | Another difficulty is to find a compound active on all subclasses of | |
| − | + | metallo-beta-lactamases and even on all the enzymes within a same | |
| − | + | subclass<sup>[6]</sup>. Indeed, despite extensive academic effort, no MBL inhibitor | |
| − | + | is yet close to the clinic<sup>[7]</sup>.The dilemma of MBLs clinical therapy has | |
| − | + | inspired us to seek for the Achilles' Heel of MBLs. | |
| − | + | </p> | |
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
| − | </section> | + | </div> |
| − | + | </section> | |
| − | + | <!-- End / Section --> | |
| − | + | <section class="awe-section" style="background: white;padding-top: 0px" id="id-3"> | |
| − | + | <div class="container"> | |
| − | + | <div class="row"> | |
| − | + | <!-- title --> | |
| − | + | <div class="title"> | |
| − | + | <h2 class="title__title">Inspirations | |
| − | + | <p class="title__text"></p> | |
| − | + | </h2> | |
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10 "> | |
| − | + | <!-- pricing-01__element --> | |
| − | + | <div class="pricing-01__element wow fadeInUp" data-wow-duration="1s" data-wow-delay="0.2s" | |
| − | + | data-wow-offset="20" data-wow-iteration="1"> | |
| − | + | <header class="pricing-01__header"> | |
| − | + | <h2 class="pricing-01__title" style="font-size: 25px;text-align: left">1.Previous iGEM | |
| − | + | teams' project related to beta-lactamases: UiOslo Norway 2016.</h2> | |
| − | + | </header> | |
| + | <div class="pricing-01__body"> | ||
<span class="pricing-01__price"> | <span class="pricing-01__price"> | ||
<p class="pricing-01__words" style="font-size: 20px;text-align: left;padding: 30px"> | <p class="pricing-01__words" style="font-size: 20px;text-align: left;padding: 30px"> | ||
| Line 1,218: | Line 1,213: | ||
</p> | </p> | ||
</span> | </span> | ||
| − | + | </div> | |
| − | + | </div><!-- End / pricing-01__element --> | |
| − | + | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10 "> | |
| − | + | <!-- pricing-01__element --> | |
| − | + | <div class="pricing-01__element wow fadeInUp" data-wow-duration="1s" data-wow-delay="0.2s" | |
| − | + | data-wow-offset="20" data-wow-iteration="1"><span></span> | |
| − | + | <header class="pricing-01__header"> | |
| − | + | <h2 class="pricing-01__title" style="font-size: 25px;text-align: left">2.Joint assays | |
| − | + | inspired by three articles</h2> | |
| − | + | </header> | |
| + | <div class="pricing-01__body"> | ||
<span class="pricing-01__price"> | <span class="pricing-01__price"> | ||
<p class="pricing-01__words" style="font-size: 20px;text-align: left;padding: 30px"> | <p class="pricing-01__words" style="font-size: 20px;text-align: left;padding: 30px"> | ||
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</p> | </p> | ||
</span> | </span> | ||
| − | + | </div> | |
| − | + | </div><!-- End / pricing-01__element --> | |
| − | + | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10 "> | |
| − | + | <!-- pricing-01__element --> | |
| − | + | <div class="pricing-01__element wow fadeInUp" data-wow-duration="1s" data-wow-delay="0.2s" | |
| − | + | data-wow-offset="20" data-wow-iteration="1"><span></span> | |
| − | + | <header class="pricing-01__header"> | |
| − | + | <h2 class="pricing-01__title" style="font-size: 25px;text-align: left">3.Advice from | |
| − | + | professors</h2> | |
| − | + | </header> | |
| + | <div class="pricing-01__body"> | ||
<span class="pricing-01__price"> | <span class="pricing-01__price"> | ||
<p class="pricing-01__words" style="font-size: 20px;text-align: left;padding: 30px"> | <p class="pricing-01__words" style="font-size: 20px;text-align: left;padding: 30px"> | ||
| Line 1,270: | Line 1,265: | ||
</p> | </p> | ||
</span> | </span> | ||
| − | + | </div> | |
| − | + | </div><!-- End / pricing-01__element --> | |
| − | + | ||
</div> | </div> | ||
</div> | </div> | ||
| − | </section> | + | </div> |
| − | + | </section> | |
| − | + | <section class="awe-section" style="background: white;padding-top: 0px" id="id-4"> | |
| − | + | <div class="container"> | |
| − | + | <div class="row"> | |
| − | + | <!-- title --> | |
| − | + | <div class="title"> | |
| − | + | <h2 class="title__title" style="color: #11253b">Methods</h2> | |
| − | + | </div><!-- End / title --> | |
| − | + | <!-- about --> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10 "> | |
| − | + | <div class="about__words"> | |
| − | + | <h2 class="fw-600 fz-30" style="color: #11253b">Build gene circuits</h2> | |
| − | + | <p style="color: black; font-size:18px;line-height: 2.1"> | |
| − | + | We use synthetic biology methods to design and construct gene circuits. Moreover, | |
| − | + | using the prokaryotic cell (E. coli) as the chassis cell. We built multiple genetic | |
| − | + | circuits that ultimately ensured that each one we used matched perfectly with the | |
| − | + | chassis, resulting in our target protein. | |
| − | + | </p> | |
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10"> | |
| − | + | <div class="about__img" style="text-align: center"> | |
| − | + | <img src="https://2019.igem.org/wiki/images/2/2f/T--TJUSLS_China--description-5.png"> | |
| − | + | </div> <!-- 1260 * 750 --> | |
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10 "> | |
| − | + | <div class="about__words"> | |
| − | + | <h2 class="fw-600 fz-30" style="color: #11253b">Special fluorescent probe CDC-1</h2> | |
| − | + | <p style="color: black; font-size:18px;line-height: 2.1"> | |
| − | + | In our project we used a special fluorescent probe CDC-1, which contains a | |
| − | + | beta-lactam ring and a fluorescent group. When the beta-lactam ring is hydrolyzed by | |
| − | + | metallo-beta-lactamases, the probe emits an easily recognizable fluorescence. It is | |
| − | + | a property that will be exploited many times in our subsequent steps. | |
| − | + | </p> | |
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10"> | |
| − | + | <div class="about__img" style="text-align: center"> | |
| − | + | <img src="https://2019.igem.org/wiki/images/7/7e/T--TJUSLS_China--description-6.png"> | |
| − | + | </div> <!-- 1260 * 750 --> | |
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10 "> | |
| − | + | <div class="about__words"> | |
| − | + | <h2 class="fw-600 fz-30" style="color: #11253b">Determination of Enzyme Activity</h2> | |
| − | + | <p style="color: black; font-size:18px;line-height: 2.1"> | |
| − | + | We used fluorescent probe CDC-1 as the substrate to determine the kinetic parameters | |
| − | + | of related enzymes and the optimal buffer system, and established four enzyme | |
| − | + | activity systems for the screening of inhibitors in the next step. | |
| − | + | </p> | |
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10"> | |
| − | + | <div class="about__img" style="text-align: center"> | |
| − | + | <img src="https://2019.igem.org/wiki/images/7/73/T--TJUSLS_China--description-7.png"> | |
| − | + | </div> <!-- 1260 * 750 --> | |
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10 "> | |
| − | + | <div class="about__words"> | |
| − | + | <h2 class="fw-600 fz-30" style="color: #11253b">Fluorescent High-throughput Screening | |
| − | + | </h2> | |
| − | + | <p style="color: black; font-size:18px;line-height: 2.1"> | |
| − | + | At the molecular level,referring to the established virtual screening model, we set up | |
| − | + | high-throughput screening system based on the | |
| − | + | fluorescent probe CDC-1, and selected effective inhibitors, in which there is a | |
| − | + | broad spectrum inhibitor, whose inhibition efficiency is more than 90% from over | |
| − | + | 4000 drugs in FDA approved libraries and traditional Chinese drug libraries. We also | |
| − | + | measured IC<sub>50</sub>value of the screened inhibitors to assess the inhibition | |
| + | ability. | ||
| + | </p> | ||
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10"> | |
| − | + | <div class="about__img" style="text-align: center"> | |
| − | + | <img src="https://2019.igem.org/wiki/images/6/6c/T--TJUSLS_China--description-8.png"> | |
| − | + | </div> <!-- 1260 * 750 --> | |
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10 "> | |
| − | + | <div class="about__words"> | |
| − | + | <h2 class="fw-600 fz-30" style="color: #11253b">Living bacterial cells detection based | |
| − | + | on UV-visible Spectroscopy</h2> | |
| − | + | <p style="color: black; font-size:18px;line-height: 2.1"> | |
| − | + | At the cellular level, we combined some of the typical beta-lactams with selected | |
| − | + | inhibitors to act on E.coli which can express target MBL, and evaluated the effects | |
| − | + | of inhibitors on living cells by measuring the absorbance curve of antibiotics. | |
| − | + | </p> | |
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
| − | + | <div class="row"> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10"> | |
| − | + | <div class="about__img" style="text-align: center"> | |
| − | + | <img src="https://2019.igem.org/wiki/images/f/f9/T--TJUSLS_China--description-9.png"> | |
| − | + | </div> <!-- 1260 * 750 --> | |
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
| − | </section> | + | </div> |
| − | + | </section> | |
| − | + | <section class="awe-section" style="background: white;padding-top: 0px;" id="id-5"> | |
| − | + | <div class="container"> | |
| − | + | <div class="row"> | |
| − | + | <!-- title --> | |
| − | + | <div class="title"> | |
| − | + | <h2 class="title__title" style="color: #11253b">Aims & Meanings</h2> | |
| − | + | </div><!-- End / title --> | |
| − | + | <!-- about --> | |
| − | + | <div class="about"> | |
| − | + | <div class="col-lg-1 "></div> | |
| − | + | <div class="col-lg-10 "> | |
| − | + | <div class="about__words"> | |
| − | + | <p style="color: black; font-size:18px;line-height: 2.1"> | |
| − | + | We want to address the problem that there are no clinically available inhibitors of | |
| − | + | beta-lactamase. We hope that we can contribute to solving issue, especially the | |
| − | + | mechanism about MBLs in the treatment of disease clinical problem. For this, we | |
| − | + | screened out broad spectrum inhibitors that are expected to be used clinically to | |
| − | + | regenerate antibiotics, which can possibly solve the current medical dilemma of | |
| − | + | drug-resistant bacteria. Besides, we tested therapeutic effect in live bacteria by | |
| − | + | combining beta-lactams with inhibitors, and proved that the inhibitors we screened | |
| − | + | were effective in preventing MBLs-carrying bacteria from hydrolyzing antibiotics. We | |
| − | </ | + | have successfully hit the Achilles' Heel of MBLs! |
| + | </p> | ||
</div> | </div> | ||
</div> | </div> | ||
</div> | </div> | ||
| − | + | </div> | |
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| − | + | <p style="color: black; font-size:18px;line-height: 2.1"> | |
| − | + | For the further research, we will use clinically collected resistant strains to test | |
| − | + | the effectiveness of our inhibitors. | |
| − | + | </p> | |
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| − | + | <p style="color: black; font-size:18px;line-height: 2.1"> | |
| − | + | For therapeutics, we consider putting the screened inhibitors into applied medicine. | |
| − | + | We also hope that with mature results, the combination of beta-lactam/beta-lactamase | |
| − | + | inhibitors therapy can establish novel medicine and save lives clinically, which | |
| − | + | will be the progress in the clinical treatment of bacterial infection. | |
| − | + | </p> | |
</div> | </div> | ||
</div> | </div> | ||
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| − | + | <h2 class="title__title" style="color: #11253b">Reference</h2> | |
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| − | + | <p style="color: black;font-size: 16px">[1] Paulo Durão, Roberto Balbontín, Isabel | |
| − | + | Gordo. Evolutionary Mechanisms Shaping the Maintenance of Antibiotic Resistance. | |
| − | + | Trends in Microbiology, Volume 26, Issue 8, 2018, pages 677-691. ISSN 0966-842X, | |
| − | + | https://doi.org/10.1016/j.tim.2018.01.005. | |
| − | + | <br>[2] Jose M. Munitaand Cesar A. Arias. Mechanisms | |
| − | + | of Antibiotic Resistance. Microbiol Spectr. 2016 Apr; 4(2): | |
| − | + | 10.1128/microbiolspec.VMBF-0016-2015. | |
| − | + | ||
| − | + | <br>[3] Caitlyn M Rotondo and Gerard D Wright. | |
| + | Inhibitors of metallo-b-lactamases. Microbiology 2017,39:96–105. | ||
<br>[4] Palzkill T. Metallo-beta-lactamase structure | <br>[4] Palzkill T. Metallo-beta-lactamase structure | ||
| − | + | and function. Ann N Y Acad Sci. 2013;1277:91–104. | |
| − | + | doi:10.1111/j.1749-6632.2012.06796.x. | |
<br>[5] Anthony R. White, Clive Kaye, James Poupard, | <br>[5] Anthony R. White, Clive Kaye, James Poupard, | ||
| − | + | Rienk Pypstra, Gary Woodnutt, Brian Wynne, Augmentin ® (amoxicillin/clavulanate) in | |
| − | + | the treatment of community-acquired respiratory tract infection: a review of the | |
| − | + | continuing development of an innovative antimicrobial agent , Journal of | |
| − | + | Antimicrobial Chemotherapy, Volume 53, Issue suppl_1, February 2004, Pages i3–i20, | |
| − | + | https://doi.org/10.1093/jac/dkh050. | |
| − | + | <br>[6] Carine Bebrone. Metallo-beta-lactamases | |
| − | + | (classification, activity, genetic organization, structure, zinc coordination) and | |
| − | + | their superfamily. Biochemical Pharmacology, Volume 74, Issue 12, 2007, Pages | |
| − | + | 1686-1701, ISSN 0006-2952, https://doi.org/10.1016/j.bcp.2007.05.021. | |
| − | + | <br>[7] Tooke CL, Hinchliffe P, Bragginton EC, et | |
| − | + | al. beta-Lactamases and beta-Lactamase Inhibitors in the 21st Century. J Mol Biol. | |
| − | + | 2019;431(18):3472–3500. doi:10.1016/j.jmb.2019.04.002. | |
| − | + | <br>[8] Fan Gao. Combination of two lead compounds | |
| − | + | for the treatment of TB based on beta-lactamase as a target. 2018. | |
<br>[9] Ying Ge, Ya-Jun Zhou, Ke-Wu Yang, Yi-Lin | <br>[9] Ying Ge, Ya-Jun Zhou, Ke-Wu Yang, Yi-Lin | ||
| − | + | Zhang, Yang Xiang and Yue-Juan Zhang. Real-time activity assays of b-lactamases in | |
| − | + | living bacterial cells: application to the inhibition of antibiotic-resistant E. | |
| − | + | coli strains. Royal society of Chemistry. 2017. | |
| − | + | <br>[10] Ke-Wu Yang, Yajun Zhou, Ying Ge and Yuejuan | |
| − | + | Zhang. Real-time activity monitoring of New Delhi metallo-b-lactamase-1 in living | |
| − | + | bacterial cells by UV-Vis spectroscopy. Royal society of Chemistry,2017. | |
| − | + | </p> | |
| − | + | ||
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Revision as of 13:12, 21 October 2019
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Overview
We focus our project on hitting Achilles’ Heel of metallo-beta-lactamases (MBLs), a vital class of beta-lactamases without available clinical inhibitors, produced by drug-resistant pathogens. We want to obtain the broad-spectrum inhibitor of MBLs and lay the foundation for clinical trials in the future.
Firstly we used synthetic biology methods to express a series of MBLs in E. coli. Besides, a virtual screening model has been built to discover the therapeutically effective compounds, in order to predict potential inhibitors. Next, effective compounds were screened via high-throughput screening with fluorescent probe (CDC-1) from FDA approved drug libraries and traditional Chinese medicine libraries. Also we assessed their inhibitory ability in living bacterial cells by UV-vis spectroscopy. Finally we have obtained a series of effective inhibitors of MBLs, including an excellent broad-spectrum inhibitor, which can prevent living resistant bacteria from hydrolyzing beta-lactam antibiotics. We hope these inhibitors can be ideal candidates for therapeutics for diseases caused by drug-resistant pathogens.
Background
Metallo-beta-lactamases result in beta-lactam resistance
The introduction of antibiotics represented one of the most important medical interventions in the history of global health, resulting in a dramatic reduction in human morbidity and mortality caused by bacterial infections[1]. However, the overuse of antibiotics has made the production of drug-resistant bacteria a global burden. The World Health Organization has named antibiotic resistance as one of the three most important public health threats of the 21st century[2].
The beta-lactams are the most successful class of antibiotic drugs but they are vulnerable to inactivation by a growing cadre of beta-lactamases that now number more than a thousand variants[3]. These enzymes cleave the amide bond of the beta-lactam ring thus inactivating the antibiotic.
Figure1: The hydrolysis of penicillin.
The Ambler system divides beta-lactamases into four classes, termed A, B, C and D. This year, we focus on class B that called metallo-beta-lactamases, or MBLs. Much different from other beta-lactamases, MBLs require zinc for activity and catalysis does not proceed via a covalent intermediate but rather through direct attack of a hydroxide ion that is stabilized by the Zinc ions in the active site[4]. MBLs confer resistance to most beta-lactam antibiotics including carbapenems(often seen as last line drugs)[3], making it a global threat to public health.
Figure2: The active site of blaNDM-1(a typical kind of MBLs).
Major limitation on relevant therapy：lack of clinical MBLs inhibitors
At present, due to the huge cost and low return of developing new antibiotics, the trend of developing new antibiotics has gradually declined in recent years. Alongside improvements to antibiotics themselves, combinations of existing beta-lactams with MBLs inhibitors represent the more feasible and common strategy to extend the usefulness of these existing beta-lactams.
Figure3：Schematic diagram of combining antibiotic and inhibitor
For other beta-lactamases, a number of these beta-lactam-beta-lactamases inhibitor combinations are approved for clinical use previously (for example, Amoxicillin-clavulanate is effective in the treatment of community-acquired respiratory infections[5] )，and many others are in clinical trials.
Unfortunately, the discovery of a clinically useful, specific inhibitor of metallo-beta-lactamases is made difficult by the fact that this compound must remain inactive towards the human proteins which are members of the metallo-beta-lactamase superfamily, or other metallo-enzymes such as the angiotensin converting enzyme. Another difficulty is to find a compound active on all subclasses of metallo-beta-lactamases and even on all the enzymes within a same subclass[6]. Indeed, despite extensive academic effort, no MBL inhibitor is yet close to the clinic[7].The dilemma of MBLs clinical therapy has inspired us to seek for the Achilles' Heel of MBLs.
Inspirations
1.Previous iGEM teams' project related to beta-lactamases: UiOslo Norway 2016.
They use beta-lactamase as a diagnostic tool through cleavage of the beta-lactam ring in Nitrocefin. When the color of patients’ urine changes, the presence of resistant bacteria will be detected. As for us, can we take the idea of diagnostic further to therapeutics?
2.Joint assays inspired by three articles
A master’s degree essay [8] inspired us to do the high-throughput screening with extent fluorescent probe. He focused on the treatment of Tuberculosis caused by beta-lactamase BlaC, and screened out some inhibitors, whose influence were verified together with antibiotics in living bacteria.
To detect whether there exists a rapid way to demonstrate the effect of inhibitors in living bacterial cells, two articles [9][10] provided us a real-time activity assay approach using UV-visible spectroscopy
3.Advice from professors
During our brainstorming, we interviewed professors who research on antimicrobial resistance and other fields. They affirmed our project and gave some useful instructions. For example, Prof. Weihui Wu suggested us to test in living bacteria, and Prof. Tao Wang suggested us to quantitate the measurement of fluorescent.
Methods
Build gene circuits
We use synthetic biology methods to design and construct gene circuits. Moreover, using the prokaryotic cell (E. coli) as the chassis cell. We built multiple genetic circuits that ultimately ensured that each one we used matched perfectly with the chassis, resulting in our target protein.
Special fluorescent probe CDC-1
In our project we used a special fluorescent probe CDC-1, which contains a beta-lactam ring and a fluorescent group. When the beta-lactam ring is hydrolyzed by metallo-beta-lactamases, the probe emits an easily recognizable fluorescence. It is a property that will be exploited many times in our subsequent steps.
Determination of Enzyme Activity
We used fluorescent probe CDC-1 as the substrate to determine the kinetic parameters of related enzymes and the optimal buffer system, and established four enzyme activity systems for the screening of inhibitors in the next step.
Fluorescent High-throughput Screening
At the molecular level,referring to the established virtual screening model, we set up high-throughput screening system based on the fluorescent probe CDC-1, and selected effective inhibitors, in which there is a broad spectrum inhibitor, whose inhibition efficiency is more than 90% from over 4000 drugs in FDA approved libraries and traditional Chinese drug libraries. We also measured IC50value of the screened inhibitors to assess the inhibition ability.
Living bacterial cells detection based on UV-visible Spectroscopy
At the cellular level, we combined some of the typical beta-lactams with selected inhibitors to act on E.coli which can express target MBL, and evaluated the effects of inhibitors on living cells by measuring the absorbance curve of antibiotics.
Aims & Meanings
We want to address the problem that there are no clinically available inhibitors of beta-lactamase. We hope that we can contribute to solving issue, especially the mechanism about MBLs in the treatment of disease clinical problem. For this, we screened out broad spectrum inhibitors that are expected to be used clinically to regenerate antibiotics, which can possibly solve the current medical dilemma of drug-resistant bacteria. Besides, we tested therapeutic effect in live bacteria by combining beta-lactams with inhibitors, and proved that the inhibitors we screened were effective in preventing MBLs-carrying bacteria from hydrolyzing antibiotics. We have successfully hit the Achilles' Heel of MBLs!
For the further research, we will use clinically collected resistant strains to test the effectiveness of our inhibitors.
For therapeutics, we consider putting the screened inhibitors into applied medicine. We also hope that with mature results, the combination of beta-lactam/beta-lactamase inhibitors therapy can establish novel medicine and save lives clinically, which will be the progress in the clinical treatment of bacterial infection.
Reference
[1] Paulo Durão, Roberto Balbontín, Isabel
Gordo. Evolutionary Mechanisms Shaping the Maintenance of Antibiotic Resistance.
Trends in Microbiology, Volume 26, Issue 8, 2018, pages 677-691. ISSN 0966-842X,
https://doi.org/10.1016/j.tim.2018.01.005.
[2] Jose M. Munitaand Cesar A. Arias. Mechanisms
of Antibiotic Resistance. Microbiol Spectr. 2016 Apr; 4(2):
10.1128/microbiolspec.VMBF-0016-2015.
[3] Caitlyn M Rotondo and Gerard D Wright.
Inhibitors of metallo-b-lactamases. Microbiology 2017,39:96–105.
[4] Palzkill T. Metallo-beta-lactamase structure
and function. Ann N Y Acad Sci. 2013;1277:91–104.
doi:10.1111/j.1749-6632.2012.06796.x.
[5] Anthony R. White, Clive Kaye, James Poupard,
Rienk Pypstra, Gary Woodnutt, Brian Wynne, Augmentin ® (amoxicillin/clavulanate) in
the treatment of community-acquired respiratory tract infection: a review of the
continuing development of an innovative antimicrobial agent , Journal of
Antimicrobial Chemotherapy, Volume 53, Issue suppl_1, February 2004, Pages i3–i20,
https://doi.org/10.1093/jac/dkh050.
[6] Carine Bebrone. Metallo-beta-lactamases
(classification, activity, genetic organization, structure, zinc coordination) and
their superfamily. Biochemical Pharmacology, Volume 74, Issue 12, 2007, Pages
1686-1701, ISSN 0006-2952, https://doi.org/10.1016/j.bcp.2007.05.021.
[7] Tooke CL, Hinchliffe P, Bragginton EC, et
al. beta-Lactamases and beta-Lactamase Inhibitors in the 21st Century. J Mol Biol.
2019;431(18):3472–3500. doi:10.1016/j.jmb.2019.04.002.
[8] Fan Gao. Combination of two lead compounds
for the treatment of TB based on beta-lactamase as a target. 2018.
[9] Ying Ge, Ya-Jun Zhou, Ke-Wu Yang, Yi-Lin
Zhang, Yang Xiang and Yue-Juan Zhang. Real-time activity assays of b-lactamases in
living bacterial cells: application to the inhibition of antibiotic-resistant E.
coli strains. Royal society of Chemistry. 2017.
[10] Ke-Wu Yang, Yajun Zhou, Ying Ge and Yuejuan
Zhang. Real-time activity monitoring of New Delhi metallo-b-lactamase-1 in living
bacterial cells by UV-Vis spectroscopy. Royal society of Chemistry,2017.
CONTACT US
EMAIL
tju2019igem@163.com
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No.92 Weijin Road, Nankai District, Tianjin, China