Nicotinamide adenine dinucleotide phosphate-specific glutamate dehydrogenase of Neurospora.
Neurospora NADP-specific glutamate dehydrogenase that was treated with iodoacetate, iodoacetamide, or N-ethylmaleimide to block the thiol groups was cleaved with cyanogen bromide. Of the expected 10 peptides, based on a methionine content of 9 residues, 8 were obtained in pure form and 2 were handled as a mixture. The fragments ranged in size from 9 to 109 residues. In addition, there were isolated 6 peptides, produced by anomalous cleavage at the carboxyl groups of tryptophan residues, and two by hydrolysis of an aspartyl-proline bond. Preliminary separation of these peptides was accomplished by gel filtration followed by either ion-exchange chromatography of the larger peptides or by paper chromatography and paper electrophoresis of the smaller fragments. Ordering of the CNBr fragments in sequence was based upon sequences of tryptic and chymotryptic peptides obtained in another laboratory. The complete sequence of the protein is presented. The amino acid sequences of the bovine and chicken liver glutamate dehydrogenases previously determined show considerable homology with the NADP-specific enzyme of Neurospora in the NH2-terminal half of the molecule; this includes the region of the specifically reactive lysine residue and the portion of the sequence that has been implicated in coenzyme binding. Particularly striking is the fact that most of the residues conserved among the three homologous proteins would be expected to be important for conformational, rather than catalytic, effects. This implies that the conformation of the Neurospora enzyme must be similar in parts of its structure to the vertebrate enzymes but undoubtedly differs in some regards.
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000626:Aminopeptidases; D001665:Binding Sites; D002268:Carboxypeptidases; D002918:Chymotrypsin; D003488:Cyanogen Bromide; D005033:Ethylmaleimide; D005969:Glutamate Dehydrogenase; D007461:Iodoacetates; D009249:NADP; D009491:Neurospora / Q000201:enzymology*; D010446:Peptide Fragments / Q000032:analysis; D011485:Protein Binding; D013045:Species Specificity; D013820:Thermolysin; D014357:Trypsin
D000596:Amino Acids; D007461:Iodoacetates; D010446:Peptide Fragments; D009249:NADP; D005969:Glutamate Dehydrogenase; D002268:Carboxypeptidases; D000626:Aminopeptidases; D002918:Chymotrypsin; D014357:Trypsin; D013820:Thermolysin; D005033:Ethylmaleimide; D003488:Cyanogen Bromide