文献类型：期刊文献

中文题名：流体剪切敏感型海洋灰绿曲霉AgugtA和AgugeA基因的克隆及功能分析

英文题名：Cloning and Analysis of AgugtAand AgugeA Genes in Shear-Sensitive Marine-Derived Fungus Aspergillus glaucus

作者：胡伟[1];李佳欣[1];李晓霞[1];蔡孟浩[1];周祥山[1];张元兴[1]

机构：[1]华东理工大学生物工程学院,生物反应器工程国家重点实验室,上海200237

年份：2015

卷号：41

期号：4

起止页码：455

中文期刊名：华东理工大学学报(自然科学版)

外文期刊名：Journal of East China University of Science and Technology

收录：CSTPCD;;Scopus;北大核心:【北大核心2014】；CSCD:【CSCD2015_2016】；

基金：国家自然科学基金项目(41306121,31270141)

语种：中文

中文关键词：海洋灰绿曲霉;剪切敏感;细胞壁合成;UgtA;UgeA

外文关键词：marine-derived Aspergillus glaucus;shear sensitive;cell wall formation;UgtA;UgeA

摘要：摘要:海洋灰绿曲霉HB1—19的高度剪切敏感性给反应器发酵造成很大困难。曲霉中的UDP-半乳糖基转移酶UgtA和UDP-葡萄糖差向异构酶UgeA对细胞壁的合成起着重要的作用。本文拟研究海洋灰绿曲霉中这两种酶在细胞壁合成中的作用及其与剪切敏感性的关系。首先,采用简并PCR与染色体步移技术克隆了海洋灰绿曲霉AgugtA和AgugeA的基因序列。然后,通过逆转录PCR确定了内含子及编码序列。AgugtA全长1377bp,有4个内含子,分别位于135~192,261~327,552~601,1047~1096bp之间,编码蛋白大小为383个氨基酸。AgugeA全长1322bp,有3个内含子,分别位于30~116,352~420和878~927bp之间,编码蛋白大小为371个氨基酸。通过进化树分析发现,AgugtA和AgugeA在灰绿曲霉及曲霉属其他菌株中有很高的同源性。通过比较海洋灰绿曲霉及模式茵构巢曲霉的AgugeA基因敲除及AgugeA蛋白抑制剂实验发现,AgugeA是海洋灰绿曲霉的必需基因,这可能与海洋灰绿曲霉的高度剪切敏感性有关。
Marine-derived filamentous fungus Aspergillus glaucus HB1-19 is sensitive to mechanical shear stress and thus causes great difficulties in bioreactor fermentation.UDP-galactofuranose transporter(UgtA)and UDP-glucose epimerase(UgeA)play critical roles in wall formation.In this work,we aim to study their functions in cell wall synthesis and also their relationships with shear-sensitivity.Degenerate PCR and genome walking were used to obtain the nucleotide sequences of AgugtA and AgugeA.AgugtA is 1377 bp containing four introns locating at 135--192,261--327,552--601,1047--1096 bp separately,and codes a protein of 383 amino acids.AgugeA is 1322 bp containing three introns locating at 30--116,352--420,878--927 bp separately,and codes a protein of 371 amino acids.Phylogenetic analysis suggested that AgugtA and AgugeA are evolutionally conserved and probably possess the same effects to their homologs in Aspergillus spp.Finally,deletion attampts of AgugeA genetically or inhibition of AgugeA at the protein level severely suppressed the growth of Aspergillus glaucus HBI-19,indicating that AguaeA is essential for HB1-19,which could be closely related with its high shear sensitivity.