A new protein folding screen: Application to the ligand binding domains of a glutamate and kainate receptor and to lysozyme and carbonic anhydrase

Neali Armstrong, Alexandre De Lencastre, Eric Gouaux

Research output: Contribution to journalArticle

96 Citations (Scopus)

Abstract

Production of folded and biologically active protein from Escherichia coli derived inclusion bodies can only be accomplished if a scheme exists for in vitro naturation. Motivated by the need for a rapid and statistically meaningful method of determining and evaluating protein folding conditions, we have designed a new fractional factorial protein ridding screen. The screen includes 12 factors shown by previous experiments to enhance protein folding and it incorporates the 12 factors into 16 different folding conditions. By examining a 1/256th fraction of the full factorial, multiple folding conditions were determined for the ligand binding domains from glutamate and kainate receptors, and for lysozyme and carbonic anhydrase B. The impact of each factor on the formation of biologically active material was estimated by calculating factor main effects. Factors and corresponding levels such as pH (8.5) and L-arginine (0.5 M) consistently had a positive effect on protein folding, whereas detergent (0.3 mM lauryl maltoside) and nonpolar additive (0.4 M sucrose) were detrimental to the folding of these four proteins. One of the 16 conditions yielded the most folded material for three out of the four proteins. Our results suggest that this protein folding screen will be generally useful in determining whether other proteins will fold in vitro and, if so, what factors are important. Furthermore, fractional factorial folding screens are well suited to the evaluation of previously untested factors on protein folding.

Original languageEnglish (US)
Pages (from-to)1475-1483
Number of pages9
JournalProtein Science
Volume8
Issue number7
StatePublished - 1999
Externally publishedYes

Fingerprint

Kainic Acid Receptors
Protein folding
Carbonic Anhydrases
Protein Folding
Glutamate Receptors
Muramidase
Ligands
Proteins
Carbonic Anhydrase I
Escherichia coli Proteins
Detergents
Inclusion Bodies
Sucrose
Arginine
Experiments

Keywords

  • Bacterial expression
  • Fractional factorial screen
  • Inclusion bodies
  • Protein folding

ASJC Scopus subject areas

  • Biochemistry

Cite this

A new protein folding screen : Application to the ligand binding domains of a glutamate and kainate receptor and to lysozyme and carbonic anhydrase. / Armstrong, Neali; De Lencastre, Alexandre; Gouaux, Eric.

In: Protein Science, Vol. 8, No. 7, 1999, p. 1475-1483.

Research output: Contribution to journalArticle

@article{83e8cd7f91c64e29bc36b115a95627d0,
title = "A new protein folding screen: Application to the ligand binding domains of a glutamate and kainate receptor and to lysozyme and carbonic anhydrase",
abstract = "Production of folded and biologically active protein from Escherichia coli derived inclusion bodies can only be accomplished if a scheme exists for in vitro naturation. Motivated by the need for a rapid and statistically meaningful method of determining and evaluating protein folding conditions, we have designed a new fractional factorial protein ridding screen. The screen includes 12 factors shown by previous experiments to enhance protein folding and it incorporates the 12 factors into 16 different folding conditions. By examining a 1/256th fraction of the full factorial, multiple folding conditions were determined for the ligand binding domains from glutamate and kainate receptors, and for lysozyme and carbonic anhydrase B. The impact of each factor on the formation of biologically active material was estimated by calculating factor main effects. Factors and corresponding levels such as pH (8.5) and L-arginine (0.5 M) consistently had a positive effect on protein folding, whereas detergent (0.3 mM lauryl maltoside) and nonpolar additive (0.4 M sucrose) were detrimental to the folding of these four proteins. One of the 16 conditions yielded the most folded material for three out of the four proteins. Our results suggest that this protein folding screen will be generally useful in determining whether other proteins will fold in vitro and, if so, what factors are important. Furthermore, fractional factorial folding screens are well suited to the evaluation of previously untested factors on protein folding.",
keywords = "Bacterial expression, Fractional factorial screen, Inclusion bodies, Protein folding",
author = "Neali Armstrong and {De Lencastre}, Alexandre and Eric Gouaux",
year = "1999",
language = "English (US)",
volume = "8",
pages = "1475--1483",
journal = "Protein Science",
issn = "0961-8368",
publisher = "Cold Spring Harbor Laboratory Press",
number = "7",

}

TY - JOUR

T1 - A new protein folding screen

T2 - Application to the ligand binding domains of a glutamate and kainate receptor and to lysozyme and carbonic anhydrase

AU - Armstrong, Neali

AU - De Lencastre, Alexandre

AU - Gouaux, Eric

PY - 1999

Y1 - 1999

N2 - Production of folded and biologically active protein from Escherichia coli derived inclusion bodies can only be accomplished if a scheme exists for in vitro naturation. Motivated by the need for a rapid and statistically meaningful method of determining and evaluating protein folding conditions, we have designed a new fractional factorial protein ridding screen. The screen includes 12 factors shown by previous experiments to enhance protein folding and it incorporates the 12 factors into 16 different folding conditions. By examining a 1/256th fraction of the full factorial, multiple folding conditions were determined for the ligand binding domains from glutamate and kainate receptors, and for lysozyme and carbonic anhydrase B. The impact of each factor on the formation of biologically active material was estimated by calculating factor main effects. Factors and corresponding levels such as pH (8.5) and L-arginine (0.5 M) consistently had a positive effect on protein folding, whereas detergent (0.3 mM lauryl maltoside) and nonpolar additive (0.4 M sucrose) were detrimental to the folding of these four proteins. One of the 16 conditions yielded the most folded material for three out of the four proteins. Our results suggest that this protein folding screen will be generally useful in determining whether other proteins will fold in vitro and, if so, what factors are important. Furthermore, fractional factorial folding screens are well suited to the evaluation of previously untested factors on protein folding.

AB - Production of folded and biologically active protein from Escherichia coli derived inclusion bodies can only be accomplished if a scheme exists for in vitro naturation. Motivated by the need for a rapid and statistically meaningful method of determining and evaluating protein folding conditions, we have designed a new fractional factorial protein ridding screen. The screen includes 12 factors shown by previous experiments to enhance protein folding and it incorporates the 12 factors into 16 different folding conditions. By examining a 1/256th fraction of the full factorial, multiple folding conditions were determined for the ligand binding domains from glutamate and kainate receptors, and for lysozyme and carbonic anhydrase B. The impact of each factor on the formation of biologically active material was estimated by calculating factor main effects. Factors and corresponding levels such as pH (8.5) and L-arginine (0.5 M) consistently had a positive effect on protein folding, whereas detergent (0.3 mM lauryl maltoside) and nonpolar additive (0.4 M sucrose) were detrimental to the folding of these four proteins. One of the 16 conditions yielded the most folded material for three out of the four proteins. Our results suggest that this protein folding screen will be generally useful in determining whether other proteins will fold in vitro and, if so, what factors are important. Furthermore, fractional factorial folding screens are well suited to the evaluation of previously untested factors on protein folding.

KW - Bacterial expression

KW - Fractional factorial screen

KW - Inclusion bodies

KW - Protein folding

UR - http://www.scopus.com/inward/record.url?scp=0032789940&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0032789940&partnerID=8YFLogxK

M3 - Article

C2 - 10422836

AN - SCOPUS:0032789940

VL - 8

SP - 1475

EP - 1483

JO - Protein Science

JF - Protein Science

SN - 0961-8368

IS - 7

ER -