Purification and characterization of an N-acylaminoacyl-peptide hydrolase from rabbit muscle.
An N-acylaminoacyl-peptide hydrolase has been purified to homogeneity (7,000-fold with 20% yield) from rabbit muscle. This overall enrichment and its general properties as a soluble protein suggest that it is of cytosolic origin and not a component of ribosomes or other cellular organelles. The enzyme has an Mr of 230,000-245,000 and a subunit Mr of 76,000-80,000. An extensive survey of the substrate specificity of the pure enzyme reveals that our earlier conclusions (Radhakrishna, G., and Wold, F. (1986) J. Biol. Chem. 261, 9572-9575) that the enzyme is specific for Ac-Met-peptides are wrong. The enzyme catalyzes the rapid removal of Ac-Thr, Ac-Ala, Ac-Met, Ac-Ser, and more slowly Ac-Gly from peptides of different lengths. Other acetylated amino acids (Cys, Tyr, Asp, Val, Phe, Ile, Leu) may be removed at 1% or less of the rate of the above good substrates from some peptide substrates. The nature of the amino acid in the second position of the acetylated peptide generally has only a minor effect on the reaction rate; however, with charged amino acids (Arg, Asp) in the second position the reaction is retarded, and with proline it is virtually abolished. Except for slow rate of hydrolysis of acetylated dipeptides, the hydrolase does not appear to be severely affected by the peptide length in the range studied (from 2 to 11 amino acid residues). The hydrolase also cleaves formylamino acids from formylated peptides. The biological function of the enzyme is not clear.
Radhakrishna
G
G
Department of Biochemistry and Molecular Biology, University of Texas Medical School, Houston 77225.
Wold
F
F
D000107:Acetylation; D000581:Amidohydrolases / Q000302:isolation & purification* / Q000378:metabolism; D000596:Amino Acids / Q000378:metabolism; D000818:Animals; D002845:Chromatography; D002850:Chromatography, Gel; D003600:Cytosol / Q000201:enzymology; D005590:Fractional Precipitation; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D009132:Muscles / Q000201:enzymology*; D011817:Rabbits; D013379:Substrate Specificity
D000596:Amino Acids; D000581:Amidohydrolases; C032846:alpha-N-acylamino acid hydrolase