作者：夏东慧,刘宁,郭秀娜,李多川
单位：山东农业大学植物保护学院,山东 泰安 271018
关键词：多糖单加氧酶,纤维素酶,C1/C4位氧化,定点突变
基金：国家科技支撑计划(2015BAD15B05),国家自然科学基金(31571949),国家高技术研究发展计划(2012AA10180402)
以嗜热毛壳菌Chaetomium thermophilum多糖单加氧酶CtPMO1为研究对象,利用薄层层析色谱法(TLC)、高效液相色谱-示差折光检测法(HPLC-RID)和飞行时间质谱(MALDI-TOF-MS)检测CtPMO1的酶活性,并根据定点突变的原理,将CtPMO1第1位的组氨酸(His1)和第166位的谷氨酰胺(Gln166)突变为H1A、Q166A和Q166E,研究两个突变位点是否参与CtPMO1的氧化作用;另外,采用3,5-二硝基水杨酸(DNS)法检测CtPMO1与纤维素酶(EGⅡ、BGLⅠ和CBHⅠ)的协同效应。研究发现CtPMO1在温度为50 ℃、pH为5.0的条件下降解磷酸膨胀纤维素(PASC),其酶解产物中不仅存在纤维二糖至纤维五糖,还存在C1氧化寡糖和C4氧化寡糖;此外,发现突变酶H1A完全丧失了酶活,Q166A丧失了C1和C4氧化活性,而Q166E保留了部分C1氧化活性;通过对CtPMO1与纤维素酶协同作用的探究,发现利用CtPMO1预处理磷酸膨胀纤维素(PASC),分别添加EGⅡ、BGLⅠ和CBHⅠ,使还原糖产量分别提高2.10倍、2.08倍和2.16倍,协同度分别是1.022、0.799和0.875。研究结果表明CtPMO1对底物具有C1和C4氧化的功能,其反应的最适温度为50 ℃、最适pH为5.0;CtPMO1活性中心氨基酸His1和平坦表面氨基酸Gln166均是关键性位点;CtPMO1预处理PASC,使纤维素酶的降解效率发生不同程度的提高。"/>template{display:none;}.mag-rich-xref-fn { cursor: pointer;}//长视频$(document).ready(function(){if($("#showLongArticleVideo") && $("#showLongArticleVideo").length && $("#showLongArticleVideo").length>0){mag_ajax_update({ele_id:'showLongArticleVideo',url:mag_currentQikanUrl() + "/CN/article/showLongArticleVideo.do?id="+$("#articleId").val()});}});//可视化图谱$(document).ready(function(){if($("#showArticle_knowledgeMap") && $("#showArticle_knowledgeMap").length && $("#showArticle_knowledgeMap").length>0){jQuery.getJSON("/Jwxb/CN/lexeme/existsCnctstInArticle.do?articleID="+$("#articleId").val(), null, function(json){if(json.result){$("#showArticle_knowledgeMap").show();}});}});$(document).ready(function(){var metaDataStr = $.trim($("#article_metaJson").val());if(metaDataStr!=''){eval("window.metaData="+metaDataStr);}if(window.metaData && (!window.metaData.authorNotesCommon_cn && !window.metaData.authorNotesCorresp_cn && window.metaData.authorNotes_cn)){window.metaData.authorNotesCommon_cn = window.metaData.authorNotes_cn;}if(window.metaData && (!window.metaData.authorNotesCommon_en && !window.metaData.authorNotesCorresp_en && window.metaData.authorNotes_en)){window.metaData.authorNotesCommon_en = window.metaData.authorNotes_en;}var _nlmdtdXml = $("#article_nlmdtdXml").val();var magDir = _nlmdtdXml.replace(/\/[^\/]+$/,'');window.metaData.magDir = magDir;new mag_vue({ el: '#metaVue', //dataUrl: mag_currentQikanUrl() + '/EN/article/getRichHtmlJson.do?articleId='+$("#articleId").val(), data:window.metaData, loading:true, doTextFun:function(text){ return doDataJsonText(text); 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来源：2022年第7期
《菌物学报》期刊编辑部
查看菌物学报杂志2022年第7期