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Nuclear magnetic relaxation studies of the conformation of adenosine 5'-triphosphate on pyruvate kinase from rabbit muscle.
J Biol Chem. 1976 Apr 25; 251(8):2412-20.JB

Abstract

The conformation of adenosine 5'-triphosphate in the manganese complex of pyruvate kinase from rabbit muscle was determined from six metal to nucleus distances derived by nuclear magnetic relaxation techniques. On the enzyme, no direct metal-ATP coordination exists. The phosphorous atoms of ATP are 4.9 to 5.1 A away from manganese, a distance which indicates either a predominantly (greater than or equal to 94%) second sphere complex or, less likely, a highly distorted inner sphere complex. Thus, water ligands or ligands from the protein might intervene between the ATP molecule and the divalent metal ion and facilitate their interaction. The metal-gammaP distance of 5 A for pyruvate kinase-bound ATP is equal to that found for the phosphorous atom of phosphoenolpyruvate and cobalt(II) on pyruvate kinase (Melamud, E., and Mildvan, A. S. (1975) J. Biol. Chem. 250, 8193-8201), which is consistent with the overlap in space of the P-enolpyruvate-phosphorus and the gammaP of ATP at the active site. This observation explains the competitive binding of these two substrates to the enzyme, as detected by NMR and by early kinetic studies. From the phosphorus data and from measurements of the relaxation rates of 3 protons of ATP in the pyruvate kinase-metal-ATP complex, the conformation of ATP was characterized as extended with distances of 6.0, 9.1, and 7.5 A from manganese to the H8, H2, and H'1 protons, respectively. The torsion angle about the glycosidic bond (chi) which defines the conformation of the enzyme-bound riboside and adenine rings was determined to be 30 degrees. In contrast, the conformation of the binary Mn(II)-ATP complex in solution is folded around the metal with direct manganese coordination of the alpha-, beta-, and gamma-phosphorus atoms, and with metal to proton distances of 4.5, 6.4, and 6.2 A for the H8, H2, and H'1 protons, suggesting a second sphere manganese-adenine interaction. The chi angle equals 90 degrees for the binary complex primarily because of the metal-base interaction. Thus, a profound change in the conformation and structure of Mn(II)-ATP from a folded chelate to an extended second sphere complex results when the nucleotide binds to pyruvate kinase.

Authors

No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

177414

Citation

Sloan, D L., and A S. Mildvan. "Nuclear Magnetic Relaxation Studies of the Conformation of Adenosine 5'-triphosphate On Pyruvate Kinase From Rabbit Muscle." The Journal of Biological Chemistry, vol. 251, no. 8, 1976, pp. 2412-20.
Sloan DL, Mildvan AS. Nuclear magnetic relaxation studies of the conformation of adenosine 5'-triphosphate on pyruvate kinase from rabbit muscle. J Biol Chem. 1976;251(8):2412-20.
Sloan, D. L., & Mildvan, A. S. (1976). Nuclear magnetic relaxation studies of the conformation of adenosine 5'-triphosphate on pyruvate kinase from rabbit muscle. The Journal of Biological Chemistry, 251(8), 2412-20.
Sloan DL, Mildvan AS. Nuclear Magnetic Relaxation Studies of the Conformation of Adenosine 5'-triphosphate On Pyruvate Kinase From Rabbit Muscle. J Biol Chem. 1976 Apr 25;251(8):2412-20. PubMed PMID: 177414.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Nuclear magnetic relaxation studies of the conformation of adenosine 5'-triphosphate on pyruvate kinase from rabbit muscle. AU - Sloan,D L, AU - Mildvan,A S, PY - 1976/4/25/pubmed PY - 1976/4/25/medline PY - 1976/4/25/entrez SP - 2412 EP - 20 JF - The Journal of biological chemistry JO - J. Biol. Chem. VL - 251 IS - 8 N2 - The conformation of adenosine 5'-triphosphate in the manganese complex of pyruvate kinase from rabbit muscle was determined from six metal to nucleus distances derived by nuclear magnetic relaxation techniques. On the enzyme, no direct metal-ATP coordination exists. The phosphorous atoms of ATP are 4.9 to 5.1 A away from manganese, a distance which indicates either a predominantly (greater than or equal to 94%) second sphere complex or, less likely, a highly distorted inner sphere complex. Thus, water ligands or ligands from the protein might intervene between the ATP molecule and the divalent metal ion and facilitate their interaction. The metal-gammaP distance of 5 A for pyruvate kinase-bound ATP is equal to that found for the phosphorous atom of phosphoenolpyruvate and cobalt(II) on pyruvate kinase (Melamud, E., and Mildvan, A. S. (1975) J. Biol. Chem. 250, 8193-8201), which is consistent with the overlap in space of the P-enolpyruvate-phosphorus and the gammaP of ATP at the active site. This observation explains the competitive binding of these two substrates to the enzyme, as detected by NMR and by early kinetic studies. From the phosphorus data and from measurements of the relaxation rates of 3 protons of ATP in the pyruvate kinase-metal-ATP complex, the conformation of ATP was characterized as extended with distances of 6.0, 9.1, and 7.5 A from manganese to the H8, H2, and H'1 protons, respectively. The torsion angle about the glycosidic bond (chi) which defines the conformation of the enzyme-bound riboside and adenine rings was determined to be 30 degrees. In contrast, the conformation of the binary Mn(II)-ATP complex in solution is folded around the metal with direct manganese coordination of the alpha-, beta-, and gamma-phosphorus atoms, and with metal to proton distances of 4.5, 6.4, and 6.2 A for the H8, H2, and H'1 protons, suggesting a second sphere manganese-adenine interaction. The chi angle equals 90 degrees for the binary complex primarily because of the metal-base interaction. Thus, a profound change in the conformation and structure of Mn(II)-ATP from a folded chelate to an extended second sphere complex results when the nucleotide binds to pyruvate kinase. SN - 0021-9258 UR - https://www.unboundmedicine.com/medline/citation/177414/Nuclear_magnetic_relaxation_studies_of_the_conformation_of_adenosine_5'_triphosphate_on_pyruvate_kinase_from_rabbit_muscle_ L2 - http://www.jbc.org/cgi/pmidlookup?view=long&pmid=177414 DB - PRIME DP - Unbound Medicine ER -