Enzymatic reduction of cobalt in cobalamin
AssigneeWisconsin Alumni Research Foundation
InventorsJorge C. Escalante-Semerena, Maris V. Fonseca
Life of this patent
Estimating the term…
Abstract
It is disclosed that the enzymes ATP:corrinoid adenosyltransferase (CobA) from Salmonella typhimurium and reduced flavodoxin (FldA) from E. coli are both necessary and sufficient, in the presence of ATP, to reduce the oxidation state of the cobalt atom in cobalamin and produce adenosylcobalamin. This makes possible the construction of plasmids to confer upon microbial fermentation hosts the ability to synthesize adenosylcobalamin, the supply of which can sometimes be a limitation on microbial growth in fermentation. Since the enzyme flavodoxin reductase (Fpr), also from E. coli, can act to reduce FldA for biological activity, that enzyme should preferably be expressed in a fermentation host as well.
Claims6
A method of producing adenosylcobalamin comprising the steps of
- providing non-adenosylated cobalamin in which the cobalt atom has been reduced to a 2+ state;
- exposing the non-adenosylated cobalamin to the bacterial enzymes ATP:corrinoid adenosyltransferase (CobA), flavodoxin (FldA), and flavodoxin reductase (Fpr) in the presence of ATP under conditions suitable for the production of adenosylcobalamin, wherein the CobA and FldA enzymes are not natively from the same host organism;
- and
- detecting adenosylcobalamin produced by the reaction.
A method of producing adenosylcobalamin comprising the steps of
- providing non-adenosylated cobalamin in which the cobalt atom has been reduced to a 2+ state;
- exposing the non-adenosylated cobalamin to the enzymes ATP:corrinoid adenosyltransferase (CobA) from Salmonella typhimurium, flavodoxin (FldA) from Escherichia coli, and flavodoxin reductase from Escherichia coli in the presence of ATP under conditions suitable for the production of adenosylcobalamin;
- and
- detecting adenosylcobalamin produced by the reaction.
A method of producing adenosylcobalamin comprising the steps of
- providing non-adenosylated cobalamin in which the cobalt atom has been reduced to a 2+ state;
- and
- exposing in vitro the non-adenosylated cobalamin to the bacterial enzymes ATP:corrinoid adenosyltransferase (CobA) from Salmonella typhimurium, flavodoxin (FldA) from Escherichia coli, and flavodoxin reductase from Escherichia coliin the presence of ATP under conditions suitable for the production of adenosylcobalamin.
Dependencies are inferred from the claim text.