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Biological and enzymatic mechanisms of polyester biodegradation by fungi.

机译：真菌对聚酯进行生物降解的生物学和酶促机制。

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摘要

This dissertation has explored the microbial and enzymatic processes that occur during the biodegradation of aliphatic polyesters by the fungus Aspergillus fumigatus. The relationship between the chemical structure of polyesters, the enzymes secreted by fungi to hydrolyze polyesters, and the enzymatic mechanisms of polyester degradation were investigated.; Naturally occurring bacterial polyesters such as poly({dollar}beta{dollar}-hydroxybutyrate) and poly({dollar}beta{dollar}-hydroxyvalerate), as well as synthetic polyesters, were found to be utilized as the sole carbon source in liquid fermentation cultures by a strain of A. fumigatus isolated from leaf compost. The enzyme secreted during fungal growth on PHB was isolated and characterized using enzyme kinetics, affinity reagents and inhibitors, SDS-PAGE, and IEF analysis to determine the biochemical characteristics of the eukaryotic PHB depolymerase.; The substrate specificity of the PHB depolymerase was evaluated using in vitro weight loss methods which determined the enzymatic rates of hydrolysis for bacterial and synthetic polyester films. The rates of polyester degradation were analyzed as a function of copolymer composition, molecular weight, and crystalline properties. Molecular modeling of polyesters suggested a relationship between polyester conformation and the susceptibility to PHB depolymerase hydrolysis.; Direct observations of enzyme-polymer interactions showed that the PHB depolymerase adsorbs strongly to PHB and other aliphatic polyesters by hydrophobic surface forces. The substrate binding behavior of the fungal PHB depolymerase was found to be non-specific, and is mediated by two independent enzyme domains.; By applying cyclic and linear PHB analogs to enzymatic hydrolysis experiments, a new understanding of the enzymatic mechanism of PHB hydrolysis was developed. Oligolide hydrolysis by the bacterial and fungal depolymerases established that the PHB depolymerases do not require free chain ends for the initiation of polymer degradation, and hydrolyze substrates in an exo mode of activity, in addition to endo chain cleavage.; Enzymes present in the supernatant of A. fumigatus grown on poly({dollar}varepsilon{dollar}-caprolactone) hydrolyzed olive oil, indicating that a lipase is also secreted by this fungus which degrades synthetic polyesters. The isolation and identification of the polyester hydrolases from A. fumigatus demonstrated that different esterase enzymes are secreted to catalyze PHB and PCL biodegradation.
机译：本文探讨了真菌烟曲霉对脂肪族聚酯进行生物降解过程中发生的微生物和酶促过程。研究了聚酯的化学结构，真菌分泌的水解聚酯的酶与聚酯降解的酶机制之间的关系。发现天然存在的细菌聚酯，例如聚（β）{羟基}-羟基丁酸酯和聚（β）{羟基}-羟基戊酸酯），以及合成聚酯，被用作液体中唯一的碳源。从叶片堆肥分离的烟曲霉菌株进行发酵培养。分离并用酶动力学，亲和试剂和抑制剂，SDS-PAGE和IEF分析来鉴定在PHB上真菌生长过程中分泌的酶，以确定真核PHB解聚酶的生化特性。使用体外失重方法评估PHB解聚酶的底物特异性，该方法确定细菌和合成聚酯薄膜的水解酶速率。分析聚酯降解速率与共聚物组成，分子量和结晶性能的关系。聚酯的分子模型表明聚酯构象与PHB解聚酶水解的敏感性之间存在关系。对酶与聚合物相互作用的直接观察表明，PHB解聚酶通过疏水表面力强烈吸附至PHB和其他脂族聚酯。发现真菌PHB解聚酶的底物结合行为是非特异性的，并且由两个独立的酶结构域介导。通过将环状和线性PHB类似物应用于酶水解实验，人们对PHB水解的酶机理有了新的认识。细菌和真菌解聚酶对寡聚内酯的水解证实，PHB解聚酶不需要游离链端来引发聚合物降解，并且除了内链裂解外，还可以以exo活性模式水解底物。在聚（{varal} varepsilon {dollar}-己内酯）水解橄榄油上生长的烟曲霉上清液中存在的酶，表明这种真菌还分泌脂肪酶，该酶降解合成聚酯。从烟曲霉中分离和鉴定聚酯水解酶表明，分泌了不同的酯酶来催化PHB和PCL生物降解。

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  • 作者

    Scherer, Thomas Marc.;

  • 作者单位

    University of Massachusetts Amherst.;

  • 授予单位 University of Massachusetts Amherst.;
  • 学科 Chemistry Polymer.; Biology Microbiology.; Chemistry Biochemistry.
  • 学位 Ph.D.
  • 年度 1996
  • 页码 310 p.
  • 总页数 310
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类 高分子化学（高聚物）;微生物学;生物化学;
  • 关键词

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