Purification of a peptidylglycine α-amidating enzyme from transplantable rat medullary thyroid carcinomas

Nozer M. Mehta, James P. Gilligan, Barry N. Jones, Arthur H. Bertelsen, Bernard A. Roos, Roger S. Birnbaum

Research output: Contribution to journalArticle

40 Citations (Scopus)

Abstract

A peptidyl glycine α-amidating activity has been isolated from total tissue extracts of rat medullary thyroid carcinoma (MTC). Purification of the activity by ammonium sulfate fractionation, Sephacryl S-300 chromatography, and strong anion-exchange chromatography at pH 6.0 has resolved at least four peaks of activity. The activity associated with peak III has been further purified to apparent homogeneity by strong anion-exchange chromatography at pH 8.0. The purified peak III enzyme has an apparent molecular mass of 75,000 Da as measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The identity of the 75,000-Da band as the α-amidating enzyme has been confirmed by recovery of activity from a nondenaturing polyacrylamide gel. The enzyme is catalytically active as a monomer, exhibits a pH optimum between 5.0 and 5.5, and has a turnover number of 300 min-1 for NdansylTyrValGly amidation at pH 5.5. The larger size, more acidic pH optimum, and higher specific activity distinguish the purified peak III rat MTC enzyme from the enzymes isolated from bovine and porcine pituitary or from frog skin.

Original languageEnglish
Pages (from-to)44-54
Number of pages11
JournalArchives of Biochemistry and Biophysics
Volume261
Issue number1
DOIs
StatePublished - Feb 15 1988
Externally publishedYes

Fingerprint

Purification
Rats
Chromatography
Enzymes
Anions
Tissue Extracts
Ammonium Sulfate
Molecular mass
Fractionation
Electrophoresis
Sodium Dodecyl Sulfate
Anura
Glycine
Polyacrylamide Gel Electrophoresis
Skin
Swine
Monomers
Medullary Thyroid cancer
Recovery
polyacrylamide gels

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Purification of a peptidylglycine α-amidating enzyme from transplantable rat medullary thyroid carcinomas. / Mehta, Nozer M.; Gilligan, James P.; Jones, Barry N.; Bertelsen, Arthur H.; Roos, Bernard A.; Birnbaum, Roger S.

In: Archives of Biochemistry and Biophysics, Vol. 261, No. 1, 15.02.1988, p. 44-54.

Research output: Contribution to journalArticle

Mehta, Nozer M. ; Gilligan, James P. ; Jones, Barry N. ; Bertelsen, Arthur H. ; Roos, Bernard A. ; Birnbaum, Roger S. / Purification of a peptidylglycine α-amidating enzyme from transplantable rat medullary thyroid carcinomas. In: Archives of Biochemistry and Biophysics. 1988 ; Vol. 261, No. 1. pp. 44-54.
@article{43f437f537c44e9784ffea063d16679e,
title = "Purification of a peptidylglycine α-amidating enzyme from transplantable rat medullary thyroid carcinomas",
abstract = "A peptidyl glycine α-amidating activity has been isolated from total tissue extracts of rat medullary thyroid carcinoma (MTC). Purification of the activity by ammonium sulfate fractionation, Sephacryl S-300 chromatography, and strong anion-exchange chromatography at pH 6.0 has resolved at least four peaks of activity. The activity associated with peak III has been further purified to apparent homogeneity by strong anion-exchange chromatography at pH 8.0. The purified peak III enzyme has an apparent molecular mass of 75,000 Da as measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The identity of the 75,000-Da band as the α-amidating enzyme has been confirmed by recovery of activity from a nondenaturing polyacrylamide gel. The enzyme is catalytically active as a monomer, exhibits a pH optimum between 5.0 and 5.5, and has a turnover number of 300 min-1 for NdansylTyrValGly amidation at pH 5.5. The larger size, more acidic pH optimum, and higher specific activity distinguish the purified peak III rat MTC enzyme from the enzymes isolated from bovine and porcine pituitary or from frog skin.",
author = "Mehta, {Nozer M.} and Gilligan, {James P.} and Jones, {Barry N.} and Bertelsen, {Arthur H.} and Roos, {Bernard A.} and Birnbaum, {Roger S.}",
year = "1988",
month = "2",
day = "15",
doi = "10.1016/0003-9861(88)90102-6",
language = "English",
volume = "261",
pages = "44--54",
journal = "Archives of Biochemistry and Biophysics",
issn = "0003-9861",
publisher = "Academic Press Inc.",
number = "1",

}

TY - JOUR

T1 - Purification of a peptidylglycine α-amidating enzyme from transplantable rat medullary thyroid carcinomas

AU - Mehta, Nozer M.

AU - Gilligan, James P.

AU - Jones, Barry N.

AU - Bertelsen, Arthur H.

AU - Roos, Bernard A.

AU - Birnbaum, Roger S.

PY - 1988/2/15

Y1 - 1988/2/15

N2 - A peptidyl glycine α-amidating activity has been isolated from total tissue extracts of rat medullary thyroid carcinoma (MTC). Purification of the activity by ammonium sulfate fractionation, Sephacryl S-300 chromatography, and strong anion-exchange chromatography at pH 6.0 has resolved at least four peaks of activity. The activity associated with peak III has been further purified to apparent homogeneity by strong anion-exchange chromatography at pH 8.0. The purified peak III enzyme has an apparent molecular mass of 75,000 Da as measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The identity of the 75,000-Da band as the α-amidating enzyme has been confirmed by recovery of activity from a nondenaturing polyacrylamide gel. The enzyme is catalytically active as a monomer, exhibits a pH optimum between 5.0 and 5.5, and has a turnover number of 300 min-1 for NdansylTyrValGly amidation at pH 5.5. The larger size, more acidic pH optimum, and higher specific activity distinguish the purified peak III rat MTC enzyme from the enzymes isolated from bovine and porcine pituitary or from frog skin.

AB - A peptidyl glycine α-amidating activity has been isolated from total tissue extracts of rat medullary thyroid carcinoma (MTC). Purification of the activity by ammonium sulfate fractionation, Sephacryl S-300 chromatography, and strong anion-exchange chromatography at pH 6.0 has resolved at least four peaks of activity. The activity associated with peak III has been further purified to apparent homogeneity by strong anion-exchange chromatography at pH 8.0. The purified peak III enzyme has an apparent molecular mass of 75,000 Da as measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The identity of the 75,000-Da band as the α-amidating enzyme has been confirmed by recovery of activity from a nondenaturing polyacrylamide gel. The enzyme is catalytically active as a monomer, exhibits a pH optimum between 5.0 and 5.5, and has a turnover number of 300 min-1 for NdansylTyrValGly amidation at pH 5.5. The larger size, more acidic pH optimum, and higher specific activity distinguish the purified peak III rat MTC enzyme from the enzymes isolated from bovine and porcine pituitary or from frog skin.

UR - http://www.scopus.com/inward/record.url?scp=0023882113&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0023882113&partnerID=8YFLogxK

U2 - 10.1016/0003-9861(88)90102-6

DO - 10.1016/0003-9861(88)90102-6

M3 - Article

C2 - 3341777

AN - SCOPUS:0023882113

VL - 261

SP - 44

EP - 54

JO - Archives of Biochemistry and Biophysics

JF - Archives of Biochemistry and Biophysics

SN - 0003-9861

IS - 1

ER -