近日來自上海交通大學微生物代謝國家重點實驗室的研究人員在新研究中揭示了頭孢菌素?；傅淖约羟蟹肿訖C制,，并證實頭孢菌素酰化酶是一個?；拿?，并具有一定的外切肽酶活性，還深入探討了有關弱催化機制與分子內(nèi)張力的相互關系,。相關研究論文在線發(fā)表在發(fā)表在美國生物化學與分子生物學會會刊《生物化學期刊》（The Journal of Biological Chemistry）上。
領導這一研究的是上海交通大學鄧子新院士,，其早年畢業(yè)于華中農(nóng)業(yè)大學,，2005年當選為中國科學院院士。鄧子新院士長期從事微生物分子生物學研究,，在重要類別抗生素生物合成基因克隆,、定位、結構功能分析,、表達和遺傳調(diào)控機制,、抗生素代謝工程與藥物創(chuàng)新、天然產(chǎn)物的生物化學與組合生物合成等方面取得了系統(tǒng)性研究進展,。
在這篇文章中博士研究生殷俊等揭示了頭孢菌素?；傅诙阶约羟惺怯捎诜肿觾?nèi)相互作用，并由N-端親和基團（Ntn）催化完成,。這一新催化模型的提出預示著在剪切過程中,，該酶必須經(jīng)歷一個很大的構象變化,，以克服距離約22?的空間障礙。結合晶體學數(shù)據(jù),，運用高分辨質譜分析技術對頭孢菌素?；缚臻g肽的研究，為酶催化的動態(tài)機制提供了重要的實驗證據(jù),。此外,，文章還揭示了頭孢菌素酰化酶是一個?；拿?，并具有一定的外切肽酶活性，深入探討了有關弱催化機制與分子內(nèi)張力的相互關系,，豐富了酶催化的理論知識,。
2011年鄧子新院士研究團隊在抗生素生物合成和分子酶學研究領域取得了一系列新成果。在今年3月發(fā)表在同一期刊的論文中,，博士研究生黃婷婷等運用現(xiàn)代分子生物學與天然產(chǎn)物化學技術,，成功克隆了具有顯著抗結核分支桿菌活性的抗生素吡啶霉素的生物合成基因簇，通過系統(tǒng)的體內(nèi)遺傳學和體外分子酶學研究闡明了該抗生素生物合成的起始機制,，證實了嵌合在非核糖體合成酶中的聚酮還原酶結構域的功能,，并推測出兩個吡啶環(huán)的生物合成模型。在此基礎上運用前體導向的生物合成技術,，獲得了三個新結構吡啶霉素類似物,，為成功利用組合生物合成技術產(chǎn)生新結構、新活性的藥物或先導化合物奠定了基礎,。 （生物谷Bioon.com）
生物谷推薦原文：
The Journal of Biological Chemistry DOI：10.1074/jbc.M111.242313
The N-terminal nucleophile serine of cephalosporin acylase executes the second autoproteolytic cleavage and Acyl-peptide hydrolysis
Jun Yin, Zixin Deng, Guoping Zhao and Xi Huang
Cephalosporin acylase precursor is translated as a single polypeptide chain and folds into a self-activating pre-protein. Activation requires two peptide bond cleavages that excise an internal spacer to form the mature αβ heterodimer. Using Q-TOF LC/MS, we located the second cleavage site between E159 and G160, and detected the corresponding 10 aa spacer G160DPPDLADQG169 of CA mutants. The site of the second cleavage depended on E159: moving E into the spacer or removing 5-10 residues from the spacer sequence resulted in shorter spacers with the cleavage at the carboxylic side of E. The mutant E159D was cleaved more slowly than the wild-type CA, as were the two mutants G160A and G160L. This allowed kinetic measurements showing that the second cleavage reaction was a first-order, intra-molecular process. Glutaryl-7-amino-cephalosporanic acid (Gl-7-ACA) is the classic substrate of CA, in which the N-terminal S170 of the β-subunit, is the nucleophile. E and D resemble glutaryl, suggesting that CA might also remove N-terminal E or D from peptides. This was indeed the case, suggesting that the N-terminal nucleophile (Ntn) also performed the second proteolytic cleavage. We also found that CA is an acyl-peptide hydrolase rather than a previously expected acyl-amino acid acylase. It only exhibited exopeptidase activity for the hydrolysis of an externally added peptide, supporting the intra-molecular interaction. We propose that the final CA activation is an intra-molecular process performed by an Ntn mechanism, during which large conformational changes in the α-subunit C-terminal region are required to bridge the gap between E159 and S170.
