The C-terminal segment is essential for maintaining the quaternary structure and enzyme activity of the nitric oxide forming nitrite reductase from Achromobacter cycloclastes

Wei Chao Chang, Jang Yi Chen, Tschining Chang, Ming Yih Liu, William J. Payne, Jean Legall, Wen Chang Chang

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

We have constructed and expressed a series of mutated nitrite reductase (NIR) mutants based on the sequence of NIR from Achromobacter cycloclastes. Deleting a pentapeptide, an undecapeptide, or a heptadecapeptide from the C-terminus of NIR resulted in a series of C-terminal deletion mutated proteins designated as NIR-5, NIR-11, and NIR-17, respectively. A C-terminally extended mutated protein, NIR+8, was also produced, which contains an extra octapeptide attached to the C-terminus of the wild-type NIR. An SDS-PAGE system using tris-tricine buffer could retain the native NIR in its trimeric form, thus offering a convenient method to check the quaternary structure of NIR analogs. By using this system it was found that NIR-5 was maintained as trimer and retained 72% of wild-type enzyme activity. However, both NIR-11 and NIR-17 behaved as monomers in the SDS-PAGE and lost all their enzyme activity. Although NIR+8 maintained its trimeric structure it was enzymatically inactive. These results clearly indicate that the C-terminal undecapeptide is essential for maintaining the quaternary structure as well as the full enzymatic activity, as expected from the X-ray crystallography studies.

Original languageEnglish
Pages (from-to)782-785
Number of pages4
JournalBiochemical and Biophysical Research Communications
Volume250
Issue number3
DOIs
Publication statusPublished - Sep 29 1998
Externally publishedYes

Fingerprint

Achromobacter cycloclastes
Nitrite Reductases
Enzyme activity
Nitric Oxide
Enzymes
Polyacrylamide Gel Electrophoresis

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

The C-terminal segment is essential for maintaining the quaternary structure and enzyme activity of the nitric oxide forming nitrite reductase from Achromobacter cycloclastes. / Chang, Wei Chao; Chen, Jang Yi; Chang, Tschining; Liu, Ming Yih; Payne, William J.; Legall, Jean; Chang, Wen Chang.

In: Biochemical and Biophysical Research Communications, Vol. 250, No. 3, 29.09.1998, p. 782-785.

Research output: Contribution to journalArticle

@article{8ece504f28a24d9cb22fad955d85b6d3,
title = "The C-terminal segment is essential for maintaining the quaternary structure and enzyme activity of the nitric oxide forming nitrite reductase from Achromobacter cycloclastes",
abstract = "We have constructed and expressed a series of mutated nitrite reductase (NIR) mutants based on the sequence of NIR from Achromobacter cycloclastes. Deleting a pentapeptide, an undecapeptide, or a heptadecapeptide from the C-terminus of NIR resulted in a series of C-terminal deletion mutated proteins designated as NIR-5, NIR-11, and NIR-17, respectively. A C-terminally extended mutated protein, NIR+8, was also produced, which contains an extra octapeptide attached to the C-terminus of the wild-type NIR. An SDS-PAGE system using tris-tricine buffer could retain the native NIR in its trimeric form, thus offering a convenient method to check the quaternary structure of NIR analogs. By using this system it was found that NIR-5 was maintained as trimer and retained 72{\%} of wild-type enzyme activity. However, both NIR-11 and NIR-17 behaved as monomers in the SDS-PAGE and lost all their enzyme activity. Although NIR+8 maintained its trimeric structure it was enzymatically inactive. These results clearly indicate that the C-terminal undecapeptide is essential for maintaining the quaternary structure as well as the full enzymatic activity, as expected from the X-ray crystallography studies.",
author = "Chang, {Wei Chao} and Chen, {Jang Yi} and Tschining Chang and Liu, {Ming Yih} and Payne, {William J.} and Jean Legall and Chang, {Wen Chang}",
year = "1998",
month = "9",
day = "29",
doi = "10.1006/bbrc.1998.9316",
language = "English",
volume = "250",
pages = "782--785",
journal = "Biochemical and Biophysical Research Communications",
issn = "0006-291X",
publisher = "Elsevier B.V.",
number = "3",

}

TY - JOUR

T1 - The C-terminal segment is essential for maintaining the quaternary structure and enzyme activity of the nitric oxide forming nitrite reductase from Achromobacter cycloclastes

AU - Chang, Wei Chao

AU - Chen, Jang Yi

AU - Chang, Tschining

AU - Liu, Ming Yih

AU - Payne, William J.

AU - Legall, Jean

AU - Chang, Wen Chang

PY - 1998/9/29

Y1 - 1998/9/29

N2 - We have constructed and expressed a series of mutated nitrite reductase (NIR) mutants based on the sequence of NIR from Achromobacter cycloclastes. Deleting a pentapeptide, an undecapeptide, or a heptadecapeptide from the C-terminus of NIR resulted in a series of C-terminal deletion mutated proteins designated as NIR-5, NIR-11, and NIR-17, respectively. A C-terminally extended mutated protein, NIR+8, was also produced, which contains an extra octapeptide attached to the C-terminus of the wild-type NIR. An SDS-PAGE system using tris-tricine buffer could retain the native NIR in its trimeric form, thus offering a convenient method to check the quaternary structure of NIR analogs. By using this system it was found that NIR-5 was maintained as trimer and retained 72% of wild-type enzyme activity. However, both NIR-11 and NIR-17 behaved as monomers in the SDS-PAGE and lost all their enzyme activity. Although NIR+8 maintained its trimeric structure it was enzymatically inactive. These results clearly indicate that the C-terminal undecapeptide is essential for maintaining the quaternary structure as well as the full enzymatic activity, as expected from the X-ray crystallography studies.

AB - We have constructed and expressed a series of mutated nitrite reductase (NIR) mutants based on the sequence of NIR from Achromobacter cycloclastes. Deleting a pentapeptide, an undecapeptide, or a heptadecapeptide from the C-terminus of NIR resulted in a series of C-terminal deletion mutated proteins designated as NIR-5, NIR-11, and NIR-17, respectively. A C-terminally extended mutated protein, NIR+8, was also produced, which contains an extra octapeptide attached to the C-terminus of the wild-type NIR. An SDS-PAGE system using tris-tricine buffer could retain the native NIR in its trimeric form, thus offering a convenient method to check the quaternary structure of NIR analogs. By using this system it was found that NIR-5 was maintained as trimer and retained 72% of wild-type enzyme activity. However, both NIR-11 and NIR-17 behaved as monomers in the SDS-PAGE and lost all their enzyme activity. Although NIR+8 maintained its trimeric structure it was enzymatically inactive. These results clearly indicate that the C-terminal undecapeptide is essential for maintaining the quaternary structure as well as the full enzymatic activity, as expected from the X-ray crystallography studies.

UR - http://www.scopus.com/inward/record.url?scp=0032578587&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0032578587&partnerID=8YFLogxK

U2 - 10.1006/bbrc.1998.9316

DO - 10.1006/bbrc.1998.9316

M3 - Article

VL - 250

SP - 782

EP - 785

JO - Biochemical and Biophysical Research Communications

JF - Biochemical and Biophysical Research Communications

SN - 0006-291X

IS - 3

ER -