Effect of selective cysteine --> alanine replacements on the catalytic functions of lysine: N6-hydroxylase.
Recombinant lysine: N6-hydroxylase, rIucD, catalyzes the conversion of L-lysine to its N6-hydroxy derivative. Re-examination of the nucleotide sequence of iucD, the gene encoding for the enzyme, has revealed a few discrepancies in the data documented in literature and the corrected version is presented. The revised nucleotide sequence predicts the presence of five cysteine residues in the primary structure of IucD. Two of these residues, cysteine 51 and cysteine 158 are alkylatable by iodoacetate in the native conformation of the protein resulting in a loss of monooxygenase activity while their replacement with alanine has no such adverse effect. Site directed mutagenesis studies have enabled an assessment of the reactivity of these cysteine residue(s) towards thiol modifying agents.
Marrone
L
L
Department of Chemistry, University of Waterloo, Canada.
Viswanatha
T
T
D015086:2,6-Dichloroindophenol / Q000378:metabolism; D000409:Alanine / Q000378:metabolism; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D003488:Cyanogen Bromide; D003545:Cysteine / Q000032:analysis / Q000378:metabolism; D004228:Dithionitrobenzoic Acid / Q000378:metabolism; D005420:Flavoproteins / Q000235:genetics / Q000378:metabolism; D007461:Iodoacetates / Q000378:metabolism; D019807:Iodoacetic Acid; D008239:Lysine / Q000378:metabolism; D013058:Mass Spectrometry; D006899:Mixed Function Oxygenases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009249:NADP / Q000378:metabolism; D010446:Peptide Fragments / Q000032:analysis; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017421:Sequence Analysis; D014357:Trypsin / Q000378:metabolism
D005420:Flavoproteins; D007461:Iodoacetates; D010446:Peptide Fragments; D011994:Recombinant Proteins; D009249:NADP; D004228:Dithionitrobenzoic Acid; D015086:2,6-Dichloroindophenol; D006899:Mixed Function Oxygenases; C047196:lysine N-epsilon hydroxylase; D014357:Trypsin; D008239:Lysine; D003545:Cysteine; D000409:Alanine; D003488:Cyanogen Bromide; D019807:Iodoacetic Acid