Abstract

Branched chain keto dehydrogenase of Pseudomonas putida is an unusual enzyme whose activity is stimulated by L-valine. The enzyme has been purified to a specific activity of 10 units/mg protein. The purified enzyme contains 4 subunis with molecular weights of 49k, 46k, 39k, and 37k. The 49k protein has been isolated and identified as lipoamide dehydrogenase. The objective of the current year will be to isolate and identify the remainding subunits. The transacylase subunit will be identified by its specific labeling with N-ethyl(2,3-14C) maleimide in the presence of 2-ketoisovalerate. The dehydrogenase (E1) subunit will be identified by its reaction with ferricyanide and the release of radioactive carbon dioxide from 2-keto(1-14C) isovalerate.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK021737-08
Application #
3227113
Study Section
(SSS)
Project Start
1979-07-01
Project End
1987-08-31
Budget Start
1986-07-01
Budget End
1987-08-31
Support Year
8
Fiscal Year
1986
Total Cost
Indirect Cost

  Institution

Name
University of Oklahoma Health Sciences Center
Department
Type
School of Medicine & Dentistry
DUNS #
937727907
City
Oklahoma City
State
OK
Country
United States
Zip Code
73117

  Publications

Hester, K L; Lehman, J; Najar, F et al. (2000) Crc is involved in catabolite repression control of the bkd operons of Pseudomonas putida and Pseudomonas aeruginosa. J Bacteriol 182:1144-9
Hester, K L; Madhusudhan, K T; Sokatch, J R (2000) Catabolite repression control by crc in 2xYT medium is mediated by posttranscriptional regulation of bkdR expression in Pseudomonas putida. J Bacteriol 182:1150-3
Madhusudhan, K T; Luo, J; Sokatch, J R (1999) In vitro transcriptional studies of the bkd operon of Pseudomonas putida: L-branched-chain amino acids and D-leucine are the inducers. J Bacteriol 181:2889-94
Madhusudhan, K T; Huang, N; Braswell, E H et al. (1997) Binding of L-branched-chain amino acids causes a conformational change in BkdR. J Bacteriol 179:276-9
Madhusudhan, K T; Hester, K L; Friend, V et al. (1997) Transcriptional activation of the bkd operon of Pseudomonas putida by BkdR. J Bacteriol 179:1992-7
Huang, N; Madhusudhan, K T; Sokatch, J R (1996) Stoichiometry of BkdR to substrate DNA in Pseudomonas putida. Biochem Biophys Res Commun 223:315-9
Hester, K; Luo, J; Burns, G et al. (1995) Purification of active E1 alpha 2 beta 2 of Pseudomonas putida branched-chain-oxoacid dehydrogenase. Eur J Biochem 233:828-36
Madhusudhan, K T; Huang, N; Sokatch, J R (1995) Characterization of BkdR-DNA binding in the expression of the bkd operon of Pseudomonas putida. J Bacteriol 177:636-41
Madhusudhan, K T; Lorenz, D; Sokatch, J R (1993) The bkdR gene of Pseudomonas putida is required for expression of the bkd operon and encodes a protein related to Lrp of Escherichia coli. J Bacteriol 175:3934-40
Luo, J; Burns, G; Sokatch, J R (1993) Construction of chromosomal recA mutants of Pseudomonas putida PpG2. Gene 136:263-6

Showing the most recent 10 out of 13 publications