DOI: 10.1128/jb.148.2.647-652.1981. Purification of a branched-chain keto acid dehydrogenase from Pseudomonas putida

Purification of a branched-chain keto acid dehydrogenase from Pseudomonas putida

10.1128/jb.148.2.647-652.1981

Crossref journal-article

American Society for Microbiology

Journal of Bacteriology (235)

Abstract

We purified branched-chain keto acid dehydrogenase to a specific activity of 10 mumol/min per mg of protein from Pseudomonas putida grown on valine. The purified enzyme was active with 2-ketoisovalerate, 2-ketoisocaproate, and 2-keto-3-methylvalerate in a ratio of 1.0:0.8:0.7 but showed no activity with either pyruvate or 2-ketoglutarate. There were four polypeptides in the purified enzyme (molecular weights, 49,000, 46,000, 39,000, and 37,000). The purified enzyme was deficient in the specific lipoamide dehydrogenase produced during growth on valine (molecular weight, 49,000). Branched-chain keto acid dehydrogenase required L-valine, oxidized nicotinamide adenine dinucleotide, coenzyme A, thiamine pyrophosphate, and magnesium chloride. A partially purified preparation catalyzed the oxidation of 2-keto-[1-14C]isovalerate to [14C]carbon dioxide, isobutyryl-coenzyme A, and reduced nicotinamide adenine dinucleotide in equimolar amounts. Both the Km and the Vmax for 2-ketoisovalerate were affected by the addition of L-valine to the assay mixture. However, only the Vmax values for oxidized nicotinamide adenine dinucleotide and coenzyme A were affected when L-valine was present. This suggested that valine acted by affecting the binding of branched-chain keto acids to subunit E1 of the complex.

Bibliography

Sokatch, J. R., McCully, V., & Roberts, C. M. (1981). Purification of a branched-chain keto acid dehydrogenase from Pseudomonas putida. Journal of Bacteriology, 148(2), 647â652.

Authors 3

J R Sokatch (first)
V McCully (additional)
C M Roberts (additional)

References 18 Referenced 54

Connelly , J. L. , D. J. Danner , and J. A. Bowden . 1968 . Branched chain a-keto acid metabolism. I. Isolation, purification, and partial characterization of bovine liver a-ketoisocaproic:a-keto-B-methylvaleric acid dehydrogenase . J. Biol. Chem. 243 : 1198 - 1203 . / J. Biol. Chem. / Branched chain a-keto acid metabolism. I. Isolation, purification, and partial characterization of bovine liver a-ketoisocaproic:a-keto-B-methylvaleric acid dehydrogenase by Connelly J. L. (1968)
Dancis , J. , J. Hutzler , and M. Levitz . 1963 . The diagnosis of maple syrup urine disease (branched chain ketoaciduria) by the in vitro study of the peripheral leucocyte . Pediatrics 32 : 234 - 238 . (10.1542/peds.32.2.234) / Pediatrics / The diagnosis of maple syrup urine disease (branched chain ketoaciduria) by the in vitro study of the peripheral leucocyte by Dancis J. (1963)
Danner , D. J. , S. K. Lemmon , J. C. Besharse , and L. J. Elsas . 1979 . Purification and characterization of branched chain a-keto acid dehydrogenase from bovine liver mitochondria . J. Biol. Chem. 254 : 5522 - 5526 . (10.1016/S0021-9258(18)50626-8) / J. Biol. Chem. / Purification and characterization of branched chain a-keto acid dehydrogenase from bovine liver mitochondria by Danner D. J. (1979)
Friedemann , T. E. , and G. E. Haugen . 1943 . Pyruvic acid. II. The determination of keto acids in blood and urine . J. Biol. Chem. 147 : 415 - 442 . / J. Biol. Chem. / Pyruvic acid. II. The determination of keto acids in blood and urine by Friedemann T. E. (1943)
Goldberg , A. L. , and T. W. Chang . 1978 . Regulation and significance of amino acid metabolism in skeletal muscle . Fed. Proc. 37 : 2301 - 2307 . / Fed. Proc. / Regulation and significance of amino acid metabolism in skeletal muscle by Goldberg A. L. (1978)
Upmann , F. , and L C . Tuttle . 1945 . A specific micromethod for the determination of acyl phosphates . J. Biol. Chem. 159 : 21 - 28 . (10.1016/S0021-9258(19)51298-4) / J. Biol. Chem. / A specific micromethod for the determination of acyl phosphates by Upmann F. (1945)
Massey , L K . , J. R. Sokatch , and R. S. Conrad . 1976 . Branched-chain amino acid catabolism in bacteria . Bacteriol. Rev. 40 : 42 - 54 . (10.1128/br.40.1.42-54.1976) / Bacteriol. Rev. / Branched-chain amino acid catabolism in bacteria by Massey L K (1976)
Martin , R. R. , V. D. Marshall , J. R. Sokatch , and L Unger . 1973 . Common enzymes of branched chain amino acid catabolism in Pseudomonas putida . J. Bacteriol. 115 : 198 - 204 . (10.1128/JB.115.1.198-204.1973) / J. Bacteriol. / Common enzymes of branched chain amino acid catabolism in Pseudomonas putida by Martin R. R. (1973)
Meister , A. 1957 . Preparation of a-keto acids . Methods Eazymol. 3 : 404 414. (10.1016/S0076-6879(57)03405-9) / Methods Eazymol. / Preparation of a-keto acids by Meister A. (1957)
Namba , Y. , K. Yoshizawa , A. Ejima , T. Hayashi , and T. Kaneda . 1969 . Coenzyme A- and nicotinamide adenine dinucleotide-dependent branched chain a-keto acid dehydrogenase. I. Purification and properties of the enzyme from Bacillus subtilis . J. Biol. Chem. 244 : 4437 - 4447 . (10.1016/S0021-9258(18)94337-1) / J. Biol. Chem. / Coenzyme A- and nicotinamide adenine dinucleotide-dependent branched chain a-keto acid dehydrogenase. I. Purification and properties of the enzyme from Bacillus subtilis by Namba Y. (1969)
Parker , P. J. , and P. J. Randle . 1978 . Partial purification and properties of branched chain 2-oxo acid dehydrogenase of ox liver . Biochem. J. 171 : 751 - 757 . (10.1042/bj1710751) / Biochem. J. / Partial purification and properties of branched chain 2-oxo acid dehydrogenase of ox liver by Parker P. J. (1978)
Pettit , F. IL , S. J. Yeaman , and L. J. Reed . 1978 . Purification and characterization of branched chain aketo acid dehydrogenase complex of bovine kidney . Proc. Natl. Acad. Sci. U.S.A. 75 : 4881 - 4885 . (10.1073/pnas.75.10.4881) / Proc. Natl. Acad. Sci. U.S.A. / Purification and characterization of branched chain aketo acid dehydrogenase complex of bovine kidney by Pettit F. IL (1978)
Reed , L. J. 1974 . Multienzyme complexes . Acc. Chem. Res. 7 : 40 - 46 . (10.1021/ar50074a002) / Acc. Chem. Res. / Multienzyme complexes by Reed L. J. (1974)
Reed L. J. and D. J. Cox. 1970. Multienzyme complexes p. 213-240. In P. D. Boyer (ed.) The enzymes vol. 1. Academic Press Inc. New York. (10.1016/S1874-6047(08)60167-5)
Reed , L. J. , and D. J. Cox . 1966 . Purification and resolution of the pyruvate dehydrogenase complex (Escherichia coli) . Methods Enzymol. 9 : 247 - 265 . (10.1016/0076-6879(66)09057-8) / Methods Enzymol. / Purification and resolution of the pyruvate dehydrogenase complex (Escherichia coli) by Reed L. J. (1966)
Roberts , C. M. , and J. R. Sokatch . 1978 . Branched chain amino acids as activators of branched chain keto acid dehydrogenase . Biochem. Biophys. Res. Commun. 82 : 828 - 833 . (10.1016/0006-291X(78)90857-4) / Biochem. Biophys. Res. Commun. / Branched chain amino acids as activators of branched chain keto acid dehydrogenase by Roberts C. M. (1978)
Rudiger , IL W . , U. Langenbeck , and H. W. Goedde . 1972 . Oxidation of branched chain a-keto acids in Streptococcus faecalis and its dependence on lipoic acid . Hoppe-Seyler's Z. Physiol. Chem. 353 : 875 - 882 . (10.1515/bchm2.1972.353.1.875) / Hoppe-Seyler's Z. Physiol. Chem. / Oxidation of branched chain a-keto acids in Streptococcus faecalis and its dependence on lipoic acid by Rudiger IL W (1972)
Sokatch , J. R. , V. McCully , J. Gebrosky , and D. J. Sokatch . 1981 . Isolation of a specific lipoamide dehydrogenase for a branched-chain keto acid dehydrogenase from Pseudomonas putida . J. Bacteriol. 148 : 639 - 646 . (10.1128/jb.148.2.639-646.1981) / J. Bacteriol. / Isolation of a specific lipoamide dehydrogenase for a branched-chain keto acid dehydrogenase from Pseudomonas putida by Sokatch J. R. (1981)

Dates

Type	When
Created	5 years, 7 months ago (Jan. 3, 2020, 12:27 p.m.)
Deposited	2 years, 10 months ago (Oct. 9, 2022, 6:11 p.m.)
Indexed	1 year, 9 months ago (Nov. 23, 2023, 10:57 a.m.)
Issued	43 years, 9 months ago (Nov. 1, 1981)
Published	43 years, 9 months ago (Nov. 1, 1981)
Published Print	43 years, 9 months ago (Nov. 1, 1981)

Funders 0

None

BibTeX

@article{Sokatch_1981, title={Purification of a branched-chain keto acid dehydrogenase from Pseudomonas putida}, volume={148}, ISSN={1098-5530}, url={http://dx.doi.org/10.1128/jb.148.2.647-652.1981}, DOI={10.1128/jb.148.2.647-652.1981}, number={2}, journal={Journal of Bacteriology}, publisher={American Society for Microbiology}, author={Sokatch, J R and McCully, V and Roberts, C M}, year={1981}, month=nov, pages={647–652} }

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