One-step production of D-p-hydroxyphenylglycine by recombinant Escherichia coli strains.
作者： Y.-P. Chao , H. Fu , T.-E. Lo , P.T. Chen , J.-J. Wang
DOI: 10.1021/BP9901163
关键词: Substrate (chemistry) 、 Nucleic acid sequence 、 Recombinant DNA 、 Dihydropyrimidinase 、 Biochemistry 、 Enterobacteriaceae 、 Molecular cloning 、 Biology 、 Agrobacterium radiobacter 、 Escherichia coli
摘要: The gene encoding D-hydantoinase from Agrobacterium radiobacter NRRL B11291 was successfully cloned by use of polymerase chain reaction. A positive clone scored, and its nucleotide sequence further analyzed. analysis deleting various lengths nucleotides the amino terminus open reading frame revealed putative regions for promoter RBS site. By highly expressing both carbamoylase, recombinant Escherichia coli strains were able to convert DL-hydroxyphenyl hydantoin (DL-HPH) D-p-hydroxyphenylglycine (D-HPG) with a conversion yield 97%, accounting productivity 5 times higher than that obtained A. B11291. Immobilizing cells kappa-carrageenan could also achieve 93%, while attained 20% within same period reaction time. These results illustrate feasibility in employing E. accomplish one-step DL-HPH D-HPG. In process improving D-HPG production, activity increased 2.57-fold but carbamoylase remained constant, which resulted only 30% increase rate. It suggests is step setting pace Since substrate insoluble, achieving sufficient agitation appears be an important issue this heterogeneous system. This view supported study on repeated cells, shows reach more 90% free can recycled six times, whereas immobilized used twice. conclusion, poor reusability due fouling gel surface.
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