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Observations on N α ‐Deacetylation of Model Amino Acids and Peptides: Distribution and Purification of a Specific N‐Acyl Amino Acid Releasing Enzyme in Rat Brain
Author(s) -
Marks Neville,
Lo EeSing,
Stern Frederic,
Danho Waleed
Publication year - 1983
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1983.tb13670.x
Subject(s) - biochemistry , enzyme , chemistry , tripeptide , amino acid , acetylation , endopeptidase , cytosol , specific activity , peptide , prolyl endopeptidase , stereochemistry , gene
Nα –Acyl amino acid releasing enzyme (NAARE), an enzyme cleaving acetylMet‐Ala at the Met‐Ala bond was purified from rat brain cytosol to apparent homogeneity by salt precipitation, gel filtration, and several steps of ion exchange. Levels of NAARE exceeded acylase measured with acetylmethionine in all brain regions and subcellular fractions examined: 60% was associated with cytosol and the remainder with debris or the crude nuclear and mitochondrial‐synaptosomal subfractions. Activity was highest in pituitary and was approximately 0.5–0.6 that of liver or kidney. The purified enzyme preferentially hydrolyzed acetyl‐methionyl peptides: K m for acetylMet‐Ala was 0.93; V max , 3.5 nmol −1 ( k cat , 1185) with pH optimum of 8.9 as compared with 8.2 for acylases measured in cytosol. The purified enzyme was devoid of acylase and common exo‐and endopeptidase contamination. Structure‐activity relationships examined with synthetic formylated or acetylated peptides indicated no significant effects for di‐ or tripeptides if the second substituent was Ala, Ser, Asn, or Thr, but the activity was reduced 0.5‐fold for Leu, a branched‐chain amino acid. No hydrolysis was observed for polypeptides with five or more residues having N‐terminal acetylated Tyr (enkephalin) or Ser (α‐melanocyte‐stimulating hormone, thymosin α1 ), supporting the notion that the enzyme plays a role only in turnover of smaller peptides formed perhaps as a result of endopeptidase cleavage of proteins or polypeptides containing acetylated Met at the N terminus.