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Overexpression, purification, and characterization of the Pseudomonas denitrificans and Salmonella typhimurium gene products in vitamin B12 biosynthesis, structural analysis of vitamin B12 intermediates, and enzymatic synthesis of factor S3
dc.contributor.advisor | Scott, A. I. | |
dc.creator | Ozaki, Shin-Ichi | |
dc.date.accessioned | 2024-02-09T20:48:13Z | |
dc.date.available | 2024-02-09T20:48:13Z | |
dc.date.issued | 1992 | |
dc.identifier.uri | https://hdl.handle.net/1969.1/DISSERTATIONS-1448425 | |
dc.description | Vita | en |
dc.description | Major subject: Chemistry | en |
dc.description.abstract | Some of the Pseudomonas denitrificans cob and the Salmonella typhimurium cbi genes involved in vitamin B12 biosynthesis were overexpressed in the heterologous E. coli system and purified. The functions of P. denitrificans cobI and S. typhimurium cbiL gene products were defined as S-adenosyl methionine dependent methyltransferases catalyzing C-20 methylation of precorrin-2 respectively. The S. typhimurium cbiA gene product was characterized as adenosyl triphosphate dependent cobyrinic acid a,c-diamide synthase. It was proved that the S. typhimurium cbiF gene product catalyzes C-16 methylation of trimethylpyrrocorphin I to afford metal-free factor S3. Zinc insertion into the macrocycle took place non-enzymatically to produce factor S3, the zinc complex of a tetramethylated derivative of uroporphyrinogen I. The structures of precorrin-2, precorrin-3, factor II, factor III (intermediates of vitamin B12 biosynthesis), trimethylpyrrocorphin I (intermediate of factor S3 biosynthesis), and factor S3 derived from various 13C-enriched isotopomers of porphobilinogens were elucidated by 13C NMR. Successive addition of the electron donating methyl groups resulted in the changes in the conjugated array of C=C and C=N bonds of the main macrocycle, which were revealed in the NMR spectra. The differences between precorrin-2 and -3 or factor II and III in their 13C NMR spectra were attributed to the electron distribution of the macrocycle and the disruption of symmetry by C-20 methylation of precorrin-2. Careful consideration of line widths and the chemical shifts of their 13C signals indicate that precorrin-2 and factor II have time-averaged structures but that precorrin-3 and factor III have relatively fixed conjugation systems. | en |
dc.format.extent | xiii, 99 leaves | en |
dc.format.medium | electronic | en |
dc.format.mimetype | application/pdf | |
dc.language.iso | eng | |
dc.rights | This thesis was part of a retrospective digitization project authorized by the Texas A&M University Libraries. Copyright remains vested with the author(s). It is the user's responsibility to secure permission from the copyright holder(s) for re-use of the work beyond the provision of Fair Use. | en |
dc.rights.uri | http://rightsstatements.org/vocab/InC/1.0/ | |
dc.subject | Major chemistry | en |
dc.subject.classification | 1992 Dissertation O99 | |
dc.subject.lcsh | Pseudomonas | en |
dc.subject.lcsh | Salmonella typhimurium | en |
dc.subject.lcsh | Biochemical genetics | en |
dc.subject.lcsh | Enzymatic analysis | en |
dc.subject.lcsh | Vitamin B12 | en |
dc.title | Overexpression, purification, and characterization of the Pseudomonas denitrificans and Salmonella typhimurium gene products in vitamin B12 biosynthesis, structural analysis of vitamin B12 intermediates, and enzymatic synthesis of factor S3 | en |
dc.type | Thesis | en |
thesis.degree.discipline | Chemistry | en |
thesis.degree.grantor | Texas A&M University | en |
thesis.degree.name | Doctor of Philosophy | en |
thesis.degree.name | Ph. D | en |
thesis.degree.level | Doctorial | en |
dc.contributor.committeeMember | Fitzpatrick, P. F. | |
dc.contributor.committeeMember | Hogg, J. L. | |
dc.contributor.committeeMember | Kelly, J. W. | |
dc.type.genre | dissertations | en |
dc.type.material | text | en |
dc.format.digitalOrigin | reformatted digital | en |
dc.publisher.digital | Texas A&M University. Libraries | |
dc.identifier.oclc | 31469132 |
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