title: The use of arginine analogues for investigating the functional organization of the arginine-binding site in lobster muscle arginine kinase. Role of the 'essential' thiol group.
abstract: 1. The nature of arginine binding to lobster arginine kinase and the extent of its possible involvement with the ;essential' thiol group of the enzyme has been investigated with some inhibitory analogues of arginine. 2. Most of the analogues inhibit competitively, although mixed inhibition may occur if the alpha-carboxy group or alpha-amino group is absent. 3. The K(i) values indicate that strength of binding depends on the length of the carbon chain (l-isoleucine>l-valine>l- alpha-aminobutyrate>l-alanine) and the integrity of the substituents on the alpha-carbon atom (l-arginine>agmatine and l-ornithine>putrescine). The guanidino group probably contributes little to substrate binding, but a positive charge near the delta-nitrogen atom appears to be important (l-ornithine>l -citrulline>l-alpha-aminobutyrate). A cyclic analogue, 2-carboxymethyl-3-oxo-2,3,5,6,7,8-hexahydro-1H-imidazo [1,2-a][1,3]diazepine-8-carboxylic acid, has a low K(i) value similar to that of an equivalent straight-chain form, suggesting that arginine probably binds in a folded configuration. 4. The aliphatic l-amino acids give enzyme difference spectra similar to that with l-arginine and the integrity of the alpha-carboxy and alpha-amino groups appears to be a minimal but not sufficient requirement for this, as l-ornithine gives an atypical difference spectrum. A difference spectrum is interpreted as indicating an enzyme conformational change. No difference spectrum was observed with methylguanidine. 5. The ability of aliphatic alpha-l-amino acids to protect against inhibition by 5,5'-dithiobis-(2-nitrobenzoic acid) is proportional to the number of atoms in the carbon chain and inversely proportional to K(i). Ornithine gives greater protection than citrulline; analogues lacking the alpha-amino groups also protect. Agmatine, lacking the alpha-carboxy group, did not protect. 6. It is concluded that it is unlikely that the ;essential' thiol group in the enzyme interacts with any part of the arginine molecule during catalysis except, possibly, the alpha-carboxyl group.
mesh_terms: D000818:Animals; D001120:Arginine / Q000031:analogs & derivatives* / Q000378:metabolism*; D001122:Arginine Kinase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D001665:Binding Sites; D004228:Dithionitrobenzoic Acid / Q000494:pharmacology; D008121:Nephropidae / Q000201:enzymology; D010770:Phosphotransferases / Q000378:metabolism*; D013053:Spectrophotometry; D013329:Structure-Activity Relationship; D013438:Sulfhydryl Compounds / Q000378:metabolism
chemical_list: D013438:Sulfhydryl Compounds; D001120:Arginine; D004228:Dithionitrobenzoic Acid; D010770:Phosphotransferases; D001122:Arginine Kinase