温度与时间对BVDV Npro 蛋白裂解活性的影响
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中图分类号：S852.653 文献标识码：A 文章编号： 1000-4440（2026）03-0598-08
，，温度与时间对BVDV Npro 蛋白裂解活性的影响[J].江苏农业学报，2026，42（3）：598-605.doi：10.3969/j.issn.1000-4440.2026.03.018
Abstract：In order to explore the autocatalytic cleavage activity of N pro protease of bovine viral diarrhea virus （BVDV）and its key regulatory factors，the recombinant fusion expression vector pET-28a-N pro -C5-EGFP containing N pro （204号 self-cleavage sitewasconstructed，andEscherichiacoliBL21（DE3）wasusedasengineringbacteria.Therecombinant protein was induced to express at different temperatures （ 16‰ ， 25‰ ， 30Y ， 374C ）and different induction times （2 h， 4h，6h，8h， 10h ）.The expression of recombinant protein was detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis （SDS-PAGE）and Western blot.The results showed that the N p= -C5-EGFP fusion proteinwith a relative molecular mass of 4.54×104 could be cleaved to produce Npro protein with a relative molecular mass of 1.85×104 underinduction conditions， and had complete autocatalytic activity.The cleavage rate showed a significant linear growthtrendwiththe extension of induction time.Withthe increaseoftemperature，thecleavage rate increased first andthen decreased，and the optimum cleavage temperature was 30‰ . TheN p=0 protein contained a potential activecenter Cys69-His49 catalytic dyadwith a spatial conformation that performed biological functions.This study not only revealed that the BVDV Npro protein has obvious autocatalyticcleavageactivityafterexpresionintheprokaryoticsystem，butalsorevealedthattheenzymaticbehaviorissygisticallyregulated bytemperatureandinductiontime.Theresultsof thisstudyprovideabasis forfurtherstudyof theselfcleavage characteristics of BVDV Npro and its subsequent use as a target for antiviral drug screening.
Key words： bovine viral diarrhea virus; Npro ；prokaryotic system；self-cleavage
牛病毒性腹泻病毒（Bovine viral diarrhea virus，BVDV）感染牛群可引发繁殖障碍及免疫抑制相关的消化系统疾病、呼吸系统疾病等复杂临床症状[1]。（剩余12659字）