WIDE VARIATION IN ANTIBIOTIC RESISTANCE PROTEINS IDENTIFIED BY FUNCTIONAL METAGE

通过功能计量鉴定的抗生素抗性蛋白的广泛变异

• 批准号: 8365808
• 负责人: STANLEY FIELDS
• 金额: $ 2.18万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-01 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8365808
• 关键词: ADP Ribose Transferases; Amino Acid Sequence; Antibiotic Resistance; Biology; Chloramphenicol; DNA Library; Dihydrofolate Reductase; Escherichia coli; Family; Funding; Fungal Genome; Genes; Gentamicins; Grant; Kanamycin; Metagenomics; Methods; Microbe; National Center for Research Resources; Peptide Sequence Determination; Pharmaceutical Preparations; Principal Investigator; Protein Family; Proteins; Research; Research Infrastructure; Resistance; Resources; Rifampicin resistance; Rifampin; Screening procedure; Soil; Source; Specificity; Tetracyclines; Trimethoprim; Trimethoprim Resistance; United States National Institutes of Health; Variant; aminoglycoside acetyltransferase; cost; insight; novel; protein function; protein structure

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Most genes for antibiotic resistance present in soil microbes remain unexplored because most environmental microbes cannot be cultured. Only recently has the identification of these genes become feasible through the use of culture-independent methods. We screened a soil metagenomic DNA library in an Escherichia coli host for genes that can confer resistance to kanamycin, gentamicin, rifampin, trimethoprim, chloramphenicol, or tetracycline. The screen revealed 41 genes that encode novel protein variants of eight protein families, including aminoglycoside acetyltransferases, rifampin ADP-ribosyltransferases, dihydrofolate reductases, and transporters. Several proteins of the same family deviate considerably from each other, yet confer comparable resistance. For example, five dihydrofolate reductases sharing at most 44% amino acid sequence identity in pairwise comparisons were equivalent in conferring trimethoprim resistance. We identified variants of aminoglycoside acetyltransferases and transporters that differ in the specificity of the drugs for which they confer resistance. We also found wide variation in protein structure. Two forms of rifampin ADP-ribosyltransferases, one twice the size of the other, were similarly effective at conferring rifampin resistance, although the short form was expressed at much lower level. Functional metagenomic screening provides insight into the large variability in antibiotic resistance protein sequences, revealing divergent variants that preserve protein function.

该副本是利用资源的众多研究子项目之一 由NIH/NCRR资助的中心赠款提供。对该子弹的主要支持 而且，副投影的主要研究员可能是其他来源提供的 包括其他NIH来源。 列出的总费用可能 代表subproject使用的中心基础架构的估计量， NCRR赠款不直接向子弹或副本人员提供的直接资金。 土壤微生物中存在的大多数抗生素耐药性基因仍未探索，因为大多数环境微生物无法培养。 直到最近，通过使用独立的方法，这些基因的鉴定才能可行。 我们在大肠杆菌宿主中筛选了一个可以赋予卡纳米霉素，庆大霉素，利福平，三甲氧苄啶，氯平苯基或四环素的基因的土壤元基因组DNA文库。 屏幕揭示了41个基因，它们编码了八个蛋白质家族的新型蛋白质变异，包括氨基糖苷乙酰基转移酶，利福平ADP-核糖基转移酶，二氢叶酸还原酶和转运蛋白。 同一家族的几种蛋白质彼此相差很大，但具有可比的抗性。 例如，在成对比较中，最多有44％的氨基酸序列身份的五个二氢叶酸还原酶在赋予甲氧苄啶抗性方面是等效的。 我们确定了氨基糖苷乙酰转移酶和转运蛋白的变体，它们在其赋予耐药性的药物的特异性方面有所不同。 我们还发现蛋白质结构的差异很大。 两种形式的利福平ADP-核糖基转移酶，一种是另一个大小的两倍，在赋予利福平耐药性方面同样有效，尽管短形式在低得多的水平上表示。 功能性宏基因组筛选可深入了解抗生素耐药性蛋白序列的较大变异性，从而揭示了保留蛋白质功能的不同变体。