CAJ | 학술논문

家蚕（Bombyx mori）黄体色突变体（lem）的幼虫体壁富含墨蝶呤（SP），SP经墨蝶呤还原酶（SPR）的催化作用合成四氢生物蝶呤（BH4）。作为芳香族氨基酸羟化酶的重要辅酶，BH4的缺乏会导致多种神经性代谢综合症。前期研究已克隆获得家蚕SPR基因（BmSpr），确定了BmSpr为lem突变体的遗传本质。本实验将重组质粒 pET-24b-BmSpr转化至 E．coli 不同菌株的感受态细胞，对 BmSpr的体外表达条件进行了优化。SDS-PAGE和Western Blot的检测结果表明BmSPR融合蛋白能够在原核表达系统中得到稳定表达，酶活性分析结果显示体外表达的重组BmSPR对其底物SP有较好的催化活性。本研究为进一步以从家蚕lem突变体资源大量提纯的SP为底物，利用原核表达BmSPR，开展体外合成 BH4的应用基础研究奠定了实验基础。
가잠（Bombyx mori）황체색돌변체（lem）적유충체벽부함묵접령（SP），SP경묵접령환원매（SPR）적최화작용합성사경생물접령（BH4）。작위방향족안기산간화매적중요보매，BH4적결핍회도치다충신경성대사종합증。전기연구이극륭획득가잠SPR기인（BmSpr），학정료BmSpr위lem돌변체적유전본질。본실험장중조질립 pET-24b-BmSpr전화지 E．coli 불동균주적감수태세포，대 BmSpr적체외표체조건진행료우화。SDS-PAGE화Western Blot적검측결과표명BmSPR융합단백능구재원핵표체계통중득도은정표체，매활성분석결과현시체외표체적중조BmSPR대기저물SP유교호적최화활성。본연구위진일보이종가잠lem돌변체자원대량제순적SP위저물，이용원핵표체BmSPR，개전체외합성 BH4적응용기출연구전정료실험기출。
Sepiapterin (SP)is contained in the integument of silkworm Bombyx mori mutant lemon (lem)in high con-centration.Sepiapterin reductase (SPR)is a key enzyme to catalyze SP to synthesize tetrahydrobiopterin (BH4).BH4 is an essential cofactor for aromatic acid hydroxylases,deficiency of which has been associated with many neuropsycho-logical disorders.We have cloned the SPR gene from B.mori (BmSpr)and made it known that BmSpr was responsible for the lem mutant phenotype.In this study,the recombinant plasmid pET-24b-BmSpr was transformed into different E. coli strains and the expression conditions of BmSpr were optimized in vitro.SDS-PAGE and western blot analyses showed that BmSpr could be expressed steadily in the optimized prokaryotic expression system.The results of enzymatic analyses indicated that BmSpr exhibited great activity to the substrate of SP.Findings in this research help to apply the expressed BmSpr to synthesize BH4 in vitro by using extracted SP from the lem mutant as substrate in the future.