食品与发酵工业2026，Vol.52Issue(1)：1-10,10.DOI:10.13995/j.cnki.11-1802/ts.042463
毕赤酵母高效表达软骨素4-O-磺基转移酶
Highly efficient expression of chondroitin 4-O-sulfotransferase in Komagataella phaffii
摘要
Abstract
Chondroitin sulfate A(CSA)is an essential component in articular cartilage repair.CSA is synthesized by the hydroxyl sulfonation of N-acetylgalactosamine(GalNAc)at position 4 in chondroitin catalyzed by chondroitin 4-O-sulfotransferase-1(C4ST-1).However,the low enzyme activity of C4ST-1 limits its catalytic efficiency,thereby hindering the industrial production of CSA.For this rea-son,this study aimed to enhance C4ST-1 enzyme activity by combining recombinant strain construction and medium optimization.Initially,based on the screening and determination of Komagataella phaffii GS115 as the chassis cell,the combination of OST1-α secretion signal peptide and SUMO Pro 3 solubility-enhancing tag was further optimized for the secretion and expression of C4ST-1,and the highest en-zyme activity of C4ST-1 was 1,889.2 U/L after shake flask fermentation.Given that previous studies found inorganic salts to inhibit C4ST-1 enzyme activity and the high cost of the existing medium,this study optimized the medium components and found that omitting the ex-pensive component,yeast nitrogen base without amino acids(YNB),resulted in a 68.4％increase in C4ST-1 enzyme activity compared to the original medium.Furthermore,the screening and optimization of carbon sources,other nitrogen sources,and biotin further increased the C4ST-1 enzyme activity.Finally,fed-batch fermentation in a 5 L fermenter for 72 hours achieved a maximum enzyme activity of 5 040.7 U/L.This study not only lays the foundation for the large-scale production of CSA,but will also provide a reference for the fer-mentation and production of sulfotransferases required for the synthesis of other glycosaminoglycans(e.g.,heparin,dermatan sulfate).关键词
软骨素4-O-磺基转移酶/毕赤酵母/信号肽/促溶标签/发酵优化Key words
chondroitin 4-O-sulfotransferase/Komagataella phaffii/signal peptide/solubility-enhancing tag/fermentation optimization引用本文复制引用
谢专,张维娇,熊海波,康振..毕赤酵母高效表达软骨素4-O-磺基转移酶[J].食品与发酵工业,2026,52(1):1-10,10.基金项目
国家重点研发计划项目(2024YFF1106300) （2024YFF1106300）
国家自然基金联合基金项目(U24A20368,32400056) （U24A20368,32400056）
江苏省合成生物基础研究中心资助项目(BK20233003) （BK20233003）
江苏省基础研究计划自然科学基金-青年基金项目(BK20241616) （BK20241616）
国家资助博士后研究人员计划项目(GZC20240607) （GZC20240607）
