Purification and characterisation of Saccharomyces cerevisiae external invertase isoforms

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• 作者：Uroš ; Andjelković ; Srdjan Pić ; urić ; Zoran Vuj&#x10d ; ić
• 关键词：Invertase ; Isoforms ; Saccharomyces cerevisiae ; Enzyme stability ; Deglycosylation ; Imobillization
• 刊名：Food Chemistry
• 出版年：2010
• 出版时间：1 June 2010
• 年：2010
• 卷：120
• 期：3
• 页码：799-804
• 全文大小：468 K

文摘

Four external invertase isoforms (EINV1, EINV2, EINV3 and EINV4) from Saccharomyces cerevisiae were highly purified by isoelectric precipitation, ethanol precipitation, ion-exchange on QAE-Sephadex and gel filtration using Sephacryl S-200. Unlike previously published procedures for external invertase purification, a specially designed step elution was applied on QAE-Sephadex which enabled the separation of four isoforms. The isoforms have the same molecular mass and catalytic properties: K_m for sucrose (25.6 mM), pH optimum (3.5–5.0) and temperature optimum (60 °C), but they exhibit significant difference in pI values, thermal stability and chemical reactivity. Deglycosylation studies showed that the observed differences between isoforms arise from posttranslational modifications. Results showed that external invertase is a mixture of at least four isoforms, but in order to improve the efficiency of food industry processes, only the most stable isoform (EINV1) should be purified and utilised. Substantially different chemical reactivity of the isoforms could be used to improve the yield of covalent immobilization procedures.