中文摘要：

目的利用在大肠杆菌中高效重组表达的0-乙酰丝氨酸（硫醇）裂解酶A（OASS-A）合成药物中间体S-苯基-L-半胱氨酸，并对合成产物进行纯度及结构鉴定。方法通过PCR扩增目的基因CysK，并连接到pET-22b（4-）载体上构建原核表达质粒，重组体转到感受态大肠杆菌B121（DE，）中进行蛋白诱导表达，重组蛋白采用Ni2＋-树脂柱亲和层析纯化，利用SDS-PAGE鉴定表达纯化蛋白，通过HPLC对合成产物纯度进行检测，应用。HNMR技术对化合物进行结构鉴定。结果成功构建了OASS-A原核表达载体，在IPrG诱导下获得了高效表达。重组酶高效合成非蛋白质氨基酸S-苯基-L-半胱氨酸。合成的化合物以晶体形式析出，采用HPLC和1HNMR技术对其结构进行了鉴定，合成产物的纯度为100％。结论利用大肠杆菌中重组表达的OASS-A能够高效合成药物中间体S-苯基-L-半胱氨酸。

英文摘要：

Objective To construct expression vector of O - acetylserine sulfhydrylase A （OASS - A）, purify and deter- mine its chemical structure. Methods Full - length Cys K gene was amplified by polymerase chain reaction （PCR） and li- gated with pET- 22b（ ＋ ） expression vector to construct prokaryotic expression plasmid. The recombinant was transformed into competent E. coli BI21 （ DE3 ） for protein expression with IPTG induction. Recombinant protein was purified with Ni2 ＋ - resin affinity chromatography and identified by SDS - PAGE. The purity and chemical structure of the synthesized compound were determined using HPLC and 1H NMR techniques. Results The prokaryotic expression vector for OASS - A was successfully constructed and fusion protein was expressed at a high level with IPTG induction. The unnatural amino acids S - phenyl - L - cysteine was effectively synthesized by recombinant OASS - A. Conclusion The pharmaceutical inter- mediate S - phenyl - L - cysteine was effectively synthesized by recombinant OASS - A from E. coli.