期刊
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
卷 110, 期 37, 页码 14906-14911出版社
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1308098110
关键词
cobalt; NMR; precorrin; tetrapyrrole; metabolism
资金
- British Biotechnology and Biological Sciences Research Council [BB/E024203/1, BB/I012079/1]
- Wellcome Trust [091163/Z/10/Z]
- Biotechnology and Biological Sciences Research Council [BB/E013007/1, BB/I012079/1, BB/E024203/1] Funding Source: researchfish
- BBSRC [BB/I012079/1, BB/E013007/1, BB/E024203/1] Funding Source: UKRI
It has been known for the past 20 years that two pathways exist in nature for the de novo biosynthesis of the coenzyme form of vitamin B-12, adenosylcobalamin, representing aerobic and anaerobic routes. In contrast to the aerobic pathway, the anaerobic route has remained enigmatic because many of its intermediates have proven technically challenging to isolate, because of their inherent instability. However, by studying the anaerobic cobalamin biosynthetic pathway in Bacillus megaterium and using homologously overproduced enzymes, it has been possible to isolate all of the intermediates between uroporphyrinogen III and cobyrinic acid. Consequently, it has been possible to detail the activities of purified cobinamide biosynthesis (Cbi) proteins CbiF, CbiG, CbiD, CbiJ, CbiET, and CbiC, as well as show the direct in vitro conversion of 5-aminolevulinic acid into cobyrinic acid using a mixture of 14 purified enzymes. This approach has resulted in the isolation of the long sought intermediates, cobalt-precorrin-6A and -6B and cobalt-precorrin-8. EPR, in particular, has proven an effective technique in following these transformations with the cobalt(II) paramagnetic electron in the d(yz) orbital, rather than the typical d(z)2. This result has allowed us to speculate that the metal ion plays an unexpected role in assisting the interconversion of pathway intermediates. By determining a function for all of the pathway enzymes, we complete the tool set for cobalamin biosynthesis and pave the way for not only enhancing cobalamin production, but also design of cobalamin derivatives through their combinatorial use and modification.
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