title: Role of the N-terminus of rat pheochromocytoma tyrosine hydroxylase in the regulation of the enzyme's activity.
abstract: Activation of rat pheochromocytoma tyrosine hydroxylase by limited tryptic proteolysis was investigated. The modifications produced upon the enzyme's structure were analyzed with the use of sodium dodecyl sulfate/polyacrylamide gel electrophoresis and tyrosine hydroxylase activity was measured all through the digestion. During the proteolysis the activity of tyrosine hydroxylase was elevated threefold at the same time as a 56-kDa tryptic fragment was formed. When the enzyme was phosphorylated, at its N-terminal region, by a kinase copurified with tyrosine hydroxylase, the major 56-kDa species did not appear to be phosphorylated on the autoradiograph, suggesting that it was derived from the native subunit by cleavage of the N-terminal of the protein. The reactivity of the 2/40/15 anti-(tyrosine hydroxylase) monoclonal antibody with the N-terminal of tyrosine hydroxylase was also investigated, using the Western-blot technique. This antibody reacted with the 62-kDa hydroxylase subunit but not with the 60-kDa tryptic fragment; the amino acid sequences of these two species showed that the 60-kDa fragment lacked the first 16 N-terminal amino acids of the native molecule. These results suggest that the N-terminal region of tyrosine hydroxylase is apparently responsible for an inhibition of the hydroxylase activity and that the first N-terminal amino acids of the hydroxylase are necessary for the recognition of the enzyme by its antibody.
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001345:Autoradiography; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010673:Pheochromocytoma / Q000201:enzymology*; D010766:Phosphorylation; D051381:Rats; D014357:Trypsin / Q000378:metabolism; D014446:Tyrosine 3-Monooxygenase / Q000032:analysis / Q000276:immunology / Q000378:metabolism*
chemical_list: D000911:Antibodies, Monoclonal; D014446:Tyrosine 3-Monooxygenase; D014357:Trypsin