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TYROSINE HYDROXYLASE OF SHEEP BRAIN: SOME CATALYTIC AND CHEMICAL PROPERTIES OF THE DETERGENT‐SOLUBILIZED, PARTIALLY PURIFIED ENZYME 1
Author(s) -
Poillon W. N.
Publication year - 1973
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.1471-4159.1973.tb07517.x
Subject(s) - tyrosine 3 monooxygenase , chemistry , tyrosine hydroxylase , enzyme , tyrosine , non competitive inhibition , specific activity , peroxidase , enzyme assay , cofactor , biochemistry , chromatography
Abstract– Detergent‐solubilized tyrosine hydroxylase from the caudate nucleus of the sheep was purified 3‐fold by affinity chromatography on 3‐iodotyrosine modified agarose. Supplementation of the standard assay with 1 mM Fe 2+ resulted in only slight stimulation of the enzymic activity. The enzyme was, however, markedly inhibited by certain complexing agents specific for either Fe 2+ or Fe 3+ . Double reciprocal plots of the kinetic data for a representative complexing agent, bathophenanthroline, showed the inhibition to be competitive with O 2 (apparent K m 0.15 mM) and noncompetitive with either l ‐tyrosine or the synthetic tetrahydropterin cofactor DMPH 4 (apparent K m 's 0.12 and 0.29 mM, respectively). The combined inhibition and kinetics studies strongly suggest that brain tyrosine hydroxylase is an iron enzyme. Furthermore, the prosthetic iron very likely participates directly in catalytic function, presumably by binding molecular oxygen. The activity of ammonium sulphate‐precipitated enzyme was found to be stimulated 2‐fold by Fe 2+ , catalase or peroxidase, while untreated enzyme was markedly less affected by these agents. Since the only ostensible difference between the two preparations was the extensive aggregation present in the former case, the change in physical state evoked by ammonium sulphate precipitation appeared to be somehow related to this peculiar property of the enzyme.