化学
糖原磷酸化酶
果糖
生物化学
丙氨酸
基质(水族馆)
立体化学
酶
生物
氨基酸
生态学
作者
Tom Verhaeghe,Margo Diricks,Dirk Aerts,Wim Soetaert,Tom Desmet
出处
期刊:Journal of Molecular Catalysis B-enzymatic
[Elsevier]
日期:2013-12-01
卷期号:96: 81-88
被引量:27
标识
DOI:10.1016/j.molcatb.2013.06.014
摘要
Sucrose phosphorylase (SP) is a promising biocatalyst for the production of special sugars and glycoconjugates, but its transglycosylation activity rarely exceeds the competing hydrolytic reaction. Knowing how specificity is controlled, would allow to optimise this activity in an efficient way by means of enzyme engineering. Therefore, in this study, a map of the acceptor site of the SP from Bifidobacterium adolescentis was created by substituting each residue by alanine and analysing the influence on the affinity for both the natural (inorganic phosphate and fructose) and alternative acceptors (d-arabitol and pyridoxine). All residues examined were found to contribute to the specificity for phosphate (Arg135, Leu343, Tyr344), fructose (Tyr132, Asp342) or both (Pro134, Tyr196, His234, Gln345). Alternative acceptors that are glycosylated rather efficiently (e.g. d-arabitol) were found to interact with the same residues as fructose, whereas poor acceptors like pyridoxine do not seem to make any specific interactions with the enzyme. Furthermore, it is shown here that SP is already optimised to outcompete water as an acceptor substrate, meaning that it will be very difficult to lower its hydrolytic activity any further. Consequently, increasing the transglycosylation activity towards alternative acceptors seems to be the best strategy, although that would probably require a drastic remodelling of the acceptor site in most cases.
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