Recombinant rabbit tryptophan hydroxylase is a substrate for cAMP-dependent protein kinase

Kent E. Vrana; Paul J. Rucker; Sean C. Kumer

doi:10.1016/0024-3205(94)00639-3

Recombinant rabbit tryptophan hydroxylase is a substrate for cAMP-dependent protein kinase

Kent E. Vrana, Paul J. Rucker, Sean C. Kumer

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32 Scopus citations

Abstract

A full-length cDNA clone for rabbit tryptophan hydroxylase (TPH) was modified and subcloned into a bacterial expression vector. Expression of this gene in the protease-deficient strain of bacteria, BL21[DE3], produced TPH immunoreactive protein which exhibited enzyme activity. Treatment of the recombinant enzyme (in bacterial extracts) with the purified catalytic subunit of the cAMP-dependent protein kinase and [y-³²P]-ATP resulted in specific phosphorylation of TPH. This expression system provides a means of generating and purifying large amounts of this important enzyme. Moreover, these experiments establish that TPH will serve as an in vitro substrate for cAMP-dependent protein kinase.

Original language	English (US)
Pages (from-to)	1045-1052
Number of pages	8
Journal	Life Sciences
Volume	55
Issue number	13
DOIs	https://doi.org/10.1016/0024-3205(94)00639-3
State	Published - 1994

All Science Journal Classification (ASJC) codes

General Biochemistry, Genetics and Molecular Biology
Pharmacology, Toxicology and Pharmaceutics(all)

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10.1016/0024-3205(94)00639-3

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title = "Recombinant rabbit tryptophan hydroxylase is a substrate for cAMP-dependent protein kinase",

abstract = "A full-length cDNA clone for rabbit tryptophan hydroxylase (TPH) was modified and subcloned into a bacterial expression vector. Expression of this gene in the protease-deficient strain of bacteria, BL21[DE3], produced TPH immunoreactive protein which exhibited enzyme activity. Treatment of the recombinant enzyme (in bacterial extracts) with the purified catalytic subunit of the cAMP-dependent protein kinase and [y-32P]-ATP resulted in specific phosphorylation of TPH. This expression system provides a means of generating and purifying large amounts of this important enzyme. Moreover, these experiments establish that TPH will serve as an in vitro substrate for cAMP-dependent protein kinase.",

author = "Vrana, {Kent E.} and Rucker, {Paul J.} and Kumer, {Sean C.}",

note = "Funding Information: The authors thank Dr. Robert Roskoski (LSU Medical Center, New Orleans, LA) for the generous gift of purified tyrosine hydroxylase and PKA catalytic subunit. We are also grateful to Dr. Sheila Vrana (Bowman Gray School of Medicine) for valuable editorial and scientific comments. This work was supported by NIH grant GM-38931 (K.E.V.).",

year = "1994",

doi = "10.1016/0024-3205(94)00639-3",

language = "English (US)",

volume = "55",

pages = "1045--1052",

journal = "Life Sciences",

issn = "0024-3205",

publisher = "Elsevier Inc.",

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TY - JOUR

T1 - Recombinant rabbit tryptophan hydroxylase is a substrate for cAMP-dependent protein kinase

AU - Vrana, Kent E.

AU - Rucker, Paul J.

AU - Kumer, Sean C.

N1 - Funding Information: The authors thank Dr. Robert Roskoski (LSU Medical Center, New Orleans, LA) for the generous gift of purified tyrosine hydroxylase and PKA catalytic subunit. We are also grateful to Dr. Sheila Vrana (Bowman Gray School of Medicine) for valuable editorial and scientific comments. This work was supported by NIH grant GM-38931 (K.E.V.).

PY - 1994

Y1 - 1994

N2 - A full-length cDNA clone for rabbit tryptophan hydroxylase (TPH) was modified and subcloned into a bacterial expression vector. Expression of this gene in the protease-deficient strain of bacteria, BL21[DE3], produced TPH immunoreactive protein which exhibited enzyme activity. Treatment of the recombinant enzyme (in bacterial extracts) with the purified catalytic subunit of the cAMP-dependent protein kinase and [y-32P]-ATP resulted in specific phosphorylation of TPH. This expression system provides a means of generating and purifying large amounts of this important enzyme. Moreover, these experiments establish that TPH will serve as an in vitro substrate for cAMP-dependent protein kinase.

AB - A full-length cDNA clone for rabbit tryptophan hydroxylase (TPH) was modified and subcloned into a bacterial expression vector. Expression of this gene in the protease-deficient strain of bacteria, BL21[DE3], produced TPH immunoreactive protein which exhibited enzyme activity. Treatment of the recombinant enzyme (in bacterial extracts) with the purified catalytic subunit of the cAMP-dependent protein kinase and [y-32P]-ATP resulted in specific phosphorylation of TPH. This expression system provides a means of generating and purifying large amounts of this important enzyme. Moreover, these experiments establish that TPH will serve as an in vitro substrate for cAMP-dependent protein kinase.

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EP - 1052

JO - Life Sciences

JF - Life Sciences

IS - 13

ER -

Recombinant rabbit tryptophan hydroxylase is a substrate for cAMP-dependent protein kinase

Abstract

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