| Summary: | 碩士 === 弘光科技大學 === 生物科技研究所 === 105 === Histidine derivatives methylation is a natural antioxidant of histidine derivatives occurring in bacteria, plants, and Mammals. The chemical structure forms is a tautomer between thiol and thione. In preservation solution, primarily in the form of thione with a stable double-bond sulfur atoms to provide Oxidation. Because the best antioxidant capacity Has been used for functional food and skin care produce. At the present, synthesis of Histidine derivatives methylation including chemical and extraction methods also Succeed development biosynthesis of enzyme catalysis. Screening of specific genes by Mycobacterium and Neurospora strain more quickly biosynthesis of Histidine derivatives methylation to replace of the chemical and extraction steps. Therefore, Synthesis step to quickly production Histidine derivatives methylation was became development goal.
In this study, one step is a novel biosynthesis process for the production Histidine derivatives methylation by whole-cell catalysis histidine derivatives. Next step are synthesis of S-adenosylmethionine by enzyme catalysed. Because the enzyme substrate specificity, accroding to reference that cloned methyltransferase gene from HK402 strain and compare with Streptomyces venezuelae, Rhodococc. erythropolis PR4 and Streptomyces lividans TK24 three strain have been screened. Result showed only gene from HK402 has been cloned and functionally expressed in E. coli [E. coli (pQE-HK402)] have biosynthetic activity to histidine derivatives, the highest yield of 35.0% was obtained. The conversion yield of the E. coli (pQE-HK402) reaction condition at pH8 buffer, Without ions metal addition that methyltransferase have the best conversion efficiency. Synthesis of S-adenosylmethionine was successfully cloned and expressed in E. coli [ E. coli(pQE-SAMec)], However we found the inhibition problem of L-histidine-N (α)-methyltransferase by S-adenosylmethionine receptor ATP , L-histidine-N (α)-methyltransferase conversion rate remained 65.5% in concerted reaction. In the future to improve Histidine derivatives methylation conversion rate, we need to resolve receptor ATP inhibition and SAM activity.
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