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Role of conserved proline residues in stabilizing tryptophan synthase alpha subunit: analysis by mutants with alanine or glycine.
Proteins. 1991; 9(2):90-8.P

Abstract

To study the role of Pro residues in the conformation and conformational stability of a protein, nine mutant alpha subunits of tryptophan synthase from Escherichia coli, in which Ala or Gly was substituted for each of six Pro residues (positions 28, 57, 62, 96, 132, and 207) that are conserved in 10 microorganisms, were constructed by means of site-directed mutagenesis. The far-ultraviolet (UV) CD spectra of five mutant alpha subunits with Ala in place of Pro were identical to the spectrum of the wild-type protein, the exception being the mutant at position 207 (P207A). CD values in the far-UV region were less negative for P207A, indicating that the Pro residue at position 207 plays a role in maintaining the intact structure of the alpha subunit. The negative CD values of the Gly mutants examined (P28G, P96G, and P132G) were also decreased. Calorimetric measurements showed that the two mutants at position 28 (P28G and P28A) gave two peaks in the excess heat capacity curve, whereas the wild type and other Pro mutants had only a single peak. The stability of each mutant protein relative to that of the wild type was about the same for P57A, less for P62A and P132A, and markedly decreased for P96A and P207A, which are substituted at less mobile positions. The changes of denaturation entropy (delta delta dS) at the denaturation temperature of the wild-type protein (54.1 degrees C at pH 9.0) were positive for P57A, P62A, and P132A, but negative for P96A, P207A, and P132G.(ABSTRACT TRUNCATED AT 250 WORDS)

Authors+Show Affiliations

Yutani K
Institute for Protein Research, Osaka University, Japan.
Hayashi S
No affiliation info available
Sugisaki Y
No affiliation info available
Ogasahara K
No affiliation info available

MeSH

AlanineCalorimetryCircular DichroismEnzyme StabilityGlycineGuanidineGuanidinesMutagenesis, Site-DirectedProlineProtein ConformationProtein DenaturationThermodynamicsTryptophan Synthase

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

2008436

Citation

Yutani, K, et al. "Role of Conserved Proline Residues in Stabilizing Tryptophan Synthase Alpha Subunit: Analysis By Mutants With Alanine or Glycine." Proteins, vol. 9, no. 2, 1991, pp. 90-8.
Yutani K, Hayashi S, Sugisaki Y, et al. Role of conserved proline residues in stabilizing tryptophan synthase alpha subunit: analysis by mutants with alanine or glycine. Proteins. 1991;9(2):90-8.
Yutani, K., Hayashi, S., Sugisaki, Y., & Ogasahara, K. (1991). Role of conserved proline residues in stabilizing tryptophan synthase alpha subunit: analysis by mutants with alanine or glycine. Proteins, 9(2), 90-8.
Yutani K, et al. Role of Conserved Proline Residues in Stabilizing Tryptophan Synthase Alpha Subunit: Analysis By Mutants With Alanine or Glycine. Proteins. 1991;9(2):90-8. PubMed PMID: 2008436.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Role of conserved proline residues in stabilizing tryptophan synthase alpha subunit: analysis by mutants with alanine or glycine. AU - Yutani,K, AU - Hayashi,S, AU - Sugisaki,Y, AU - Ogasahara,K, PY - 1991/1/1/pubmed PY - 1991/1/1/medline PY - 1991/1/1/entrez SP - 90 EP - 8 JF - Proteins JO - Proteins VL - 9 IS - 2 N2 - To study the role of Pro residues in the conformation and conformational stability of a protein, nine mutant alpha subunits of tryptophan synthase from Escherichia coli, in which Ala or Gly was substituted for each of six Pro residues (positions 28, 57, 62, 96, 132, and 207) that are conserved in 10 microorganisms, were constructed by means of site-directed mutagenesis. The far-ultraviolet (UV) CD spectra of five mutant alpha subunits with Ala in place of Pro were identical to the spectrum of the wild-type protein, the exception being the mutant at position 207 (P207A). CD values in the far-UV region were less negative for P207A, indicating that the Pro residue at position 207 plays a role in maintaining the intact structure of the alpha subunit. The negative CD values of the Gly mutants examined (P28G, P96G, and P132G) were also decreased. Calorimetric measurements showed that the two mutants at position 28 (P28G and P28A) gave two peaks in the excess heat capacity curve, whereas the wild type and other Pro mutants had only a single peak. The stability of each mutant protein relative to that of the wild type was about the same for P57A, less for P62A and P132A, and markedly decreased for P96A and P207A, which are substituted at less mobile positions. The changes of denaturation entropy (delta delta dS) at the denaturation temperature of the wild-type protein (54.1 degrees C at pH 9.0) were positive for P57A, P62A, and P132A, but negative for P96A, P207A, and P132G.(ABSTRACT TRUNCATED AT 250 WORDS) SN - 0887-3585 UR - https://www.unboundmedicine.com/medline/citation/2008436/Role_of_conserved_proline_residues_in_stabilizing_tryptophan_synthase_alpha_subunit:_analysis_by_mutants_with_alanine_or_glycine_ DB - PRIME DP - Unbound Medicine ER -