(1) The two peptide fragments derived from the front and the tail of rabbit muscle myokinase possess remarkable and specific ligand-binding properties. By systematic syntheses and measurements of their equilibrium substrate binding of these two sets of peptides which encompass the binding sites (a) for MgATP2- and MgADP- and (b) for ADP3- and AMP2- of rabbit muscle myokinase, it is hoped to chemically outline and deduce the requirements for binding of the substrates to ATP-AMP transphosphorylase. One synthetic peptide (11-45, residues 1-45 of rabbit muscle myokinase) is currently under study by NMR techniques to further delineate its ligand interactions. As a concluding phase, attempts will be made to synthesize enzymatically active fragment(s). (2) To provide information on the basic process of ATP-transphosphorylation, a comparative physicochemical approach is being continued on several enzymes which have in common the ability to catalyze the transfer of a phosphoryl group, or pyrophosphoryl group, of ATP to a suitable acceptor, but which differ markedly in their substrate specificity requirements for the acceptor. These studies include the following series of enzymes: adenylate kinase isoenzymes from rabbit and calf; nucleoside diphosphokinase from yeast; creatine kinase isoenzymes from rabbit and calf; thiamin pyrophosphokinase from porcine brain. The total amino acid sequence of the rabbit and calf muscle adenylate kinases and of rabbit muscle creatine kinase have recently been reported. Efforts are now underway (a) to prepare crystals of creatine kinase suitable for X-ray analysis, and also (b) to prepare a fragment of 88 residues which may encompass the ATP-binding site of creatine kinase. (3) With the exciting discovery of an aberrant adenylate kinase from Duchenne dystrophic serum, our objectives were accordingly extended to include a detailed study of the human isoenzymes of adenylate kinase (and of creatine kinase) from normal and abnormal tissues. It is hoped that these studies may shed light on the genetics of Duchenne dystrophy. Recently the aberrant adenylate kinase has been isolated from Duchenne dystrophic serum, and efforts are now underway to deduce its primary structure as well as that of the normal human liver-type adenylate kinase, to which it bears a resemblance.
| Kuby, S A; Fleming, G; Alber, T et al. (1991) Studies on yeast nucleoside triphosphate-nucleoside diphosphate transphosphorylase (nucleoside diphosphokinase). IV. Steady-state kinetic properties with thymidine nucleotides (including 3'-azido-3'-deoxythymidine analogues). Enzyme 45:1-13 |
| Kurokawa, Y; Takenaka, H; Sumida, M et al. (1990) Multiforms of mammalian adenylate kinase and its monoclonal antibody against AK1. Enzyme 43:57-71 |
| Kuby, S A; Hamada, M; Johnson, M S et al. (1989) Studies on adenosine triphosphate transphosphorylases. XVIII. Synthesis and preparation of peptides and peptide fragments of rabbit muscle ATP-AMP transphosphorylase (adenylate kinase) and their nucleotide-binding properties. J Protein Chem 8:549-62 |
| Fry, D C; Byler, D M; Susi, H et al. (1988) Solution structure of the 45-residue MgATP-binding peptide of adenylate kinase as examined by 2-D NMR, FTIR, and CD spectroscopy. Biochemistry 27:3588-98 |
