Coenzyme B12-dependent 2-methyleneglutarate mutase from Clostridium barkeri. Protection by the substrate from inactivation by light.

Abstract:

:Partially purified 2-methyleneglutarate mutase from Clostridium barkeri was separated from 3-methylitaconate delta-isomerase by treatment with 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) followed by FPLC on the anion exchange column Mono Q in the presence of the detergent. When purified in the dark, the active mutase contained a corrinoid, most probably coenzyme B12. The enzyme was inactivated by light at the same wavelength (lambda less than 620 nm) and rate as free coenzyme B12. The rate was not influenced by oxygen or by temperature (0-37 degrees C). Reactivation of up to 50% of the original activity was achieved by incubation with coenzyme B12 and dithiothreitol. The substrates 2-methyleneglutarate (up to 40 mM) or (R)-3-methylitaconate specifically protected the enzyme from inactivation by visible light. This effect was enhanced 3-fold by raising the temperature from 0 degrees C to 37 degrees C. The data indicate that during catalysis, the Co-C bond of the coenzyme is cleaved and cannot be affected any more by light.

journal_name

FEBS Lett

journal_title

FEBS letters

authors

Michel C,Buckel W

doi

10.1016/0014-5793(91)80370-i

subject

Has Abstract

pub_date

1991-04-09 00:00:00

pages

108-10

issue

1-2

eissn

0014-5793

issn

1873-3468

pii

0014-5793(91)80370-I

journal_volume

281

pub_type

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