Nucleotide and AP sub 5 A complexes of porcine adenylate kinase: A sup 1 H and sup 19 F NMR study
- Max Planck Institute for Medical Research, Heidelberg (West Germany)
Proton and fluorine nuclear magnetic resonance spectroscopies (NMR) were used as methods to investigate binary complexes between porcine adenylate kinase (AK{sub 1}) and its substrates. The authors also studied the interaction of fluorinated substrate analogues and the supposed bisubstrate analogue P{sup 1},P{sup 5}-bis(5{prime}-adenosyl) pentaphosphate (AP{sub 5}A) with AK{sub 1} in the presence of Mg{sup 2+}. The chemical shifts of the C8-H, C2-H, and ribose C1{prime}-H resonances of both adenosine units in stoichiometric complexes of AK{sub 1} with AP{sub 5}A in the presence of Mg{sup 2+} could be determined. The C2-H resonance of one of the adenine bases experiences a downfield shift of about 0.8 ppm on binding to the enzyme. {sup 19}F NMR chemical shifts of 9-(3-fluoro-3-deoxy-{beta}-D-xylofuranosyl)adenine triphosphate (3{prime}-F-X-ATP)-Mg{sup 2+} and 9-(3-fluoro-3-deoxy-{beta}-D-xylofuranosyl)adenine monophosphate (3{prime}-F-X-AMP) bound to porcine adenylate kinase could be determined. The different chemical shifts of the bound nucleotides suggest that their mode of binding is different. Free and bound 3{prime}-F-X-ATP are in fast exchange with respect to their {sup 19}F chemical shifts, whereas free and bound 3{prime}-F-X-ATP are in slow exchange on the NMR time scale in the absence as well as in the presence of Mg{sup 2+}. This information could be used to determine the apparent dissociation constants of the nucleotides and the 3{prime}-F-X analogues in the binary complexes. A strong nuclear Overhauser effect (NOE) could be detected between one of the two C2 protons of AP{sub 5}A and the C2 proton of the imidazole ring of His{sup 36}. This proton thus belongs to the site which was earlier suggested to be the ATP{center dot}Mg{sup 2+} site for the pig as well as for the rabbit enzyme.
- OSTI ID:
- 5352196
- Journal Information:
- Biochemistry; (USA), Journal Name: Biochemistry; (USA) Vol. 28:10; ISSN 0006-2960; ISSN BICHA
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
62 RADIOLOGY AND NUCLEAR MEDICINE
ANIMALS
ATP
BARYONS
BIOCHEMICAL REACTION KINETICS
CHEMICAL SHIFT
DOMESTIC ANIMALS
ELEMENTARY PARTICLES
ENZYMES
ESTERS
FERMIONS
FLUORINE 19
FLUORINE ISOTOPES
HADRONS
ISOTOPES
KINETICS
LIGHT NUCLEI
MAGNETIC RESONANCE
MAMMALS
NMR SPECTRA
NUCLEAR MAGNETIC RESONANCE
NUCLEI
NUCLEONS
NUCLEOTIDES
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
ORGANIC PHOSPHORUS COMPOUNDS
OVERHAUSER EFFECT
PHOSPHONIC ACID ESTERS
PHOSPHORUS-GROUP TRANSFERASES
PHOSPHOTRANSFERASES
PROTONS
REACTION KINETICS
RESONANCE
SPECTRA
STABLE ISOTOPES
SWINE
TRANSFERASES
VERTEBRATES