Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1983 Apr;80(8):2226–2230. doi: 10.1073/pnas.80.8.2226

Evidence for a multiple random coupling mechanism in the alpha-ketoglutarate dehydrogenase multienzyme complex of Escherichia coli: a computer model analysis.

M L Hackert, R M Oliver, L J Reed
PMCID: PMC393791  PMID: 6403946

Abstract

A computer modeling system was used to analyze experimental data for inactivation of the Escherichia coli alpha-ketoglutarate dehydrogenase complex accompanying release of lipoic acid residues by lipoamidase and by trypsin [Stepp, L. R., Bleile, D. M., McRorie, D. K., Pettit, F. H. & Reed, L. J. (1981) Biochemistry 20, 4555-4560]. The results provide insight into the active-site coupling mechanism in the alpha-ketoglutarate dehydrogenase complex. The model studies indicate that the overall activity of the alpha-ketoglutarate dehydrogenase complex is influenced by redundancies and random processes that we describe as a multiple random coupling mechanism. More than one lipoyl moiety services each E1 subunit (alpha-ketoglutarate dehydrogenase, EC 1.2.4.2), and an extensive lipoyl-lipoyl interaction network for exchange of electrons and possibly acyl groups must also be present. The best fit between computed and experimental data was obtained with a model that has two lipoyl moieties servicing each E1 subunit and a lipoyl-lipoyl interaction network that links all lipoyl moieties on the E2 cube (dihydrolipoamide succinyltransferase, EC 2.3.1.61). The single lipoyl moiety on an E2 subunit is assumed to service the coenzyme A-dependent succinyltransferase site of that E2 subunit as well as an E3 subunit (dihydrolipoamide dehydrogenase, EC 1.6.4.3) if the latter is bound to that particular E2 subunit.

Full text

PDF
2226

Images in this article

2229Image
on p.2229

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Akiyama S. K., Hammes G. G. Elementary steps in the reaction mechanism of the pyruvate dehydrogenase multienzyme complex from Escherichia coli: kinetics of acetylation and deacetylation. Biochemistry. 1980 Sep 2;19(18):4208–4213. doi: 10.1021/bi00559a011. [DOI] [PubMed] [Google Scholar]
  2. Ambrose M. C., Perham R. N. Spin-label study of the mobility of enzyme-bound lipoic acid in the pyruvate dehydrogenase multienzyme complex of Escherichia coli. Biochem J. 1976 May 1;155(2):429–432. doi: 10.1042/bj1550429. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Angelides K. J., Hammes G. G. Fluorescence studies of the pyruvate dehydrogenase multienzyme complex from Escherichia coli. Biochemistry. 1979 Apr 3;18(7):1223–1229. doi: 10.1021/bi00574a017. [DOI] [PubMed] [Google Scholar]
  4. Angelides K. J., Hammes G. G. Structural and mechanistic studies of the alpha-ketoglutarate dehydrogenase multienzyme complex from Escherichia coli. Biochemistry. 1979 Dec 11;18(25):5531–5537. doi: 10.1021/bi00592a001. [DOI] [PubMed] [Google Scholar]
  5. Bates D. L., Danson M. J., Hale G., Hooper E. A., Perham R. N. Self-assembly and catalytic activity of the pyruvate dehydrogenase multienzyme complex of Escherichia coli. Nature. 1977 Jul 28;268(5618):313–316. doi: 10.1038/268313a0. [DOI] [PubMed] [Google Scholar]
  6. Bleile D. M., Munk P., Oliver R. M., Reed L. J. Subunit structure of dihydrolipoyl transacetylase component of pyruvate dehydrogenase complex from Escherichia coli. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4385–4389. doi: 10.1073/pnas.76.9.4385. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Cate R. L., Roche T. E., Davis L. C. Rapid intersite transfer of acetyl groups and movement of pyruvate dehydrogenase component in the kidney pyruvate dehydrogenase complex. J Biol Chem. 1980 Aug 25;255(16):7556–7562. [PubMed] [Google Scholar]
  8. Coggins J. R., Hooper E. A., Perham R. N. Use of dimethyl suberimidate and novel periodate-cleavable bis(imido esters) to study the quaternary structure of the pyruvate dehydrogenase multienzyme complex of Escherichia coli. Biochemistry. 1976 Jun 15;15(12):2527–2533. doi: 10.1021/bi00657a006. [DOI] [PubMed] [Google Scholar]
  9. Collins J. H., Reed L. J. Acyl group and electron pair relay system: a network of interacting lipoyl moieties in the pyruvate and alpha-ketoglutarate dehydrogenase complexes from Escherichia coli. Proc Natl Acad Sci U S A. 1977 Oct;74(10):4223–4227. doi: 10.1073/pnas.74.10.4223. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Danson M. J., Fersht A. R., Perham R. N. Rapid intramolecular coupling of active sites in the pyruvate dehydrogenase complex of Escherichia coli: mechanism for rate enhancement in a multimeric structure. Proc Natl Acad Sci U S A. 1978 Nov;75(11):5386–5390. doi: 10.1073/pnas.75.11.5386. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Danson M. J., Perham R. N. Evidence for two lipoic acid residues per lipoate acetyltransferase chain in the pyruvate dehydrogenase multienzyme complex of Escherichia coli. Biochem J. 1976 Dec 1;159(3):677–682. doi: 10.1042/bj1590677. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Derosier D. J., Oliver R. M., Reed L. J. Crystallization and preliminary structural analysis of dihydrolipoyl transsuccinylase, the core of the 2-oxoglutarate dehydrogenase complex. Proc Natl Acad Sci U S A. 1971 Jun;68(6):1135–1137. doi: 10.1073/pnas.68.6.1135. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Fuller C. C., Reed L. J., Oliver R. M., Hackert M. L. Crystallization of a dihydrolipoyl transacetylase--dihydrolipoyl dehydrogenase subcomplex and its implications regarding the subunit structure of the pyruvate dehydrogenase complex from Escherichia coli. Biochem Biophys Res Commun. 1979 Sep 27;90(2):431–438. doi: 10.1016/0006-291x(79)91253-1. [DOI] [PubMed] [Google Scholar]
  14. Grande H. J., Van Telgen H. J., Veeger C. Symmetry and asymmetry of the pyruvate dehydrogenase complexes from Azotobacter vinelandii and Escherichia coli as reflected by fluorescence and spin-label studies. Eur J Biochem. 1976 Dec 11;71(2):509–518. doi: 10.1111/j.1432-1033.1976.tb11139.x. [DOI] [PubMed] [Google Scholar]
  15. Hale G., Perham R. N. Limited proteolysis of the pyruvate dehydrogenase multienzyme complex of Escherichia coli. Eur J Biochem. 1979 Feb 15;94(1):119–126. doi: 10.1111/j.1432-1033.1979.tb12878.x. [DOI] [PubMed] [Google Scholar]
  16. Hammes G. G. Processing of intermediates in multienzyme complexes. Biochem Soc Symp. 1981;(46):73–90. [PubMed] [Google Scholar]
  17. KOIKE M., REED L. J., CARROLL W. R. alpha-Keto acid dehydrogenation complexes. IV. Resolution and reconstitution of the Escherichia coli pyruvate dehydrogenation complex. J Biol Chem. 1963 Jan;238:30–39. [PubMed] [Google Scholar]
  18. Packman L. C., Perham R. N., Roberts G. C. Cross-linking and 1H n.m.r. spectroscopy of the pyruvate dehydrogenase complex of Escherichia coli. Biochem J. 1982 Aug 1;205(2):389–396. doi: 10.1042/bj2050389. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Perham R. N., Duckworth H. W., Roberts G. C. Mobility of polypeptide chain in the pyruvate dehydrogenase complex revealed by proton NMR. Nature. 1981 Jul 30;292(5822):474–477. doi: 10.1038/292474a0. [DOI] [PubMed] [Google Scholar]
  20. Perham R. N., Roberts G. C. Limited proteolysis and proton n.m.r. spectroscopy of the 2-oxoglutarate dehydrogenase multienzyme complex of Escherichia coli. Biochem J. 1981 Dec 1;199(3):733–740. doi: 10.1042/bj1990733. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Pettit F. H., Hamilton L., Munk P., Namihira G., Eley M. H., Willms C. R., Reed L. J. Alpha-keto acid dehydrogenase complexes. XIX. Subunit structure of the Escherichia coli alpha-ketoglutarate dehydrogenase complex. J Biol Chem. 1973 Aug 10;248(15):5282–5290. [PubMed] [Google Scholar]
  22. Reed L. J., Oliver R. M. The multienzyme alpha-keto acid dehydrogenase complexes. Brookhaven Symp Biol. 1968 Jun;21(2):397–412. [PubMed] [Google Scholar]
  23. Reed L. J., Pettit F. H., Eley M. H., Hamilton L., Collins J. H., Oliver R. M. Reconstitution of the Escherichia coli pyruvate dehydrogenase complex. Proc Natl Acad Sci U S A. 1975 Aug;72(8):3068–3072. doi: 10.1073/pnas.72.8.3068. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. SUZUKI K., REED L. J. LIPOAMIDASE. J Biol Chem. 1963 Dec;238:4021–4025. [PubMed] [Google Scholar]
  25. Shepherd G. B., Hammes G. G. Fluorescence energy transfer measurements in the pyruvate dehydrogenase multienzyme complex from Escherichia coli with chemically modified lipoic acid. Biochemistry. 1977 Nov 29;16(24):5234–5241. doi: 10.1021/bi00643a012. [DOI] [PubMed] [Google Scholar]
  26. Speckhard D. C., Ikeda B. H., Wong S. S., Frey P. A. Acetylation stoichiometry of Escherichia coli pyruvate dehydrogenase complex. Biochem Biophys Res Commun. 1977 Jul 25;77(2):708–713. doi: 10.1016/s0006-291x(77)80036-3. [DOI] [PubMed] [Google Scholar]
  27. Stepp L. R., Bleile D. M., McRorie D. K., Pettit F. H., Reed L. J. Use of trypsin and lipoamidase to study the role of lipoic acid moieties in the pyruvate and alpha-ketoglutarate dehydrogenase complexes of Escherichia coli. Biochemistry. 1981 Aug 4;20(16):4555–4560. doi: 10.1021/bi00519a007. [DOI] [PubMed] [Google Scholar]
  28. Waskiewicz D. E., Hammes G. G. Fluorescence polarization study of the alpha-ketoglutarate dehydrogenase complex from Escherichia coli. Biochemistry. 1982 Dec 7;21(25):6489–6496. doi: 10.1021/bi00268a026. [DOI] [PubMed] [Google Scholar]
  29. White R. H., Bleile D. M., Reed L. J. Lipoic acid content of dihydrolipoyl transacylases determined by isotope dilution analysis. Biochem Biophys Res Commun. 1980 May 14;94(1):78–84. doi: 10.1016/s0006-291x(80)80190-2. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES