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Tryptophan-containing alpha-subunits of the Escherichia coli tryptophan synthase. Enzymatic and urea stability properties.
J Biol Chem. 1995 Jul 28; 270(30):17712-5.JB

Abstract

Early studies suggested that the Escherichia coli tryptophan synthase alpha-subunit unfolded in a two-step process in which there was a stable intermediate composed of a native alpha-1 folding unit (residues 1-188) and a completely unfolded alpha-2 folding unit (residues 189-268). More recent evidence has indicated that such a structure for the intermediate seems unlikely. In this report, single Trp residues (absent in the wild-type alpha-subunit) are substituted separately for Phe residues at positions 139 (in alpha-1) and 258 (in alpha-2) to produce the F139W, F258W, and F139W/F258W mutant alpha-subunits. The UV absorbance and fluorescence properties of the F139W/F258W double mutant are identical with those of equimolar mixtures of the single mutants, suggesting that the Trp residue at each position can independently report the behavior of its respective folding unit. Each mutant alpha-subunit is wild-type enzymatically, and when UV absorbance is monitored, the urea-induced unfolding of the three tryptophan-containing alpha-subunits is virtually identical to the wild-type protein. These wild-type properties make these proteins attractive candidates for a fluorescence examination of the behavior of the individual folding units and the structure of potential intermediate(s) and as host proteins for the insertion of our existing destabilizing and/or stabilizing mutational alterations.

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Authors+Show Affiliations

Choi SG
Department of Biological Sciences, University of Alabama, Tuscaloosa 35487, USA.
O'Donnell SE
No affiliation info available
Sarken KD
No affiliation info available
Hardman JK
No affiliation info available

MeSH

Base SequenceEnzyme StabilityEscherichia coliMolecular Sequence DataOligodeoxyribonucleotidesProtein FoldingThermodynamicsTryptophanTryptophan SynthaseUrea

Pub Type(s)

Journal Article

Language

eng

PubMed ID

7629069

Citation

Choi, S G., et al. "Tryptophan-containing Alpha-subunits of the Escherichia Coli Tryptophan Synthase. Enzymatic and Urea Stability Properties." The Journal of Biological Chemistry, vol. 270, no. 30, 1995, pp. 17712-5.
Choi SG, O'Donnell SE, Sarken KD, et al. Tryptophan-containing alpha-subunits of the Escherichia coli tryptophan synthase. Enzymatic and urea stability properties. J Biol Chem. 1995;270(30):17712-5.
Choi, S. G., O'Donnell, S. E., Sarken, K. D., & Hardman, J. K. (1995). Tryptophan-containing alpha-subunits of the Escherichia coli tryptophan synthase. Enzymatic and urea stability properties. The Journal of Biological Chemistry, 270(30), 17712-5.
Choi SG, et al. Tryptophan-containing Alpha-subunits of the Escherichia Coli Tryptophan Synthase. Enzymatic and Urea Stability Properties. J Biol Chem. 1995 Jul 28;270(30):17712-5. PubMed PMID: 7629069.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Tryptophan-containing alpha-subunits of the Escherichia coli tryptophan synthase. Enzymatic and urea stability properties. AU - Choi,S G, AU - O'Donnell,S E, AU - Sarken,K D, AU - Hardman,J K, PY - 1995/7/28/pubmed PY - 1995/7/28/medline PY - 1995/7/28/entrez SP - 17712 EP - 5 JF - The Journal of biological chemistry JO - J Biol Chem VL - 270 IS - 30 N2 - Early studies suggested that the Escherichia coli tryptophan synthase alpha-subunit unfolded in a two-step process in which there was a stable intermediate composed of a native alpha-1 folding unit (residues 1-188) and a completely unfolded alpha-2 folding unit (residues 189-268). More recent evidence has indicated that such a structure for the intermediate seems unlikely. In this report, single Trp residues (absent in the wild-type alpha-subunit) are substituted separately for Phe residues at positions 139 (in alpha-1) and 258 (in alpha-2) to produce the F139W, F258W, and F139W/F258W mutant alpha-subunits. The UV absorbance and fluorescence properties of the F139W/F258W double mutant are identical with those of equimolar mixtures of the single mutants, suggesting that the Trp residue at each position can independently report the behavior of its respective folding unit. Each mutant alpha-subunit is wild-type enzymatically, and when UV absorbance is monitored, the urea-induced unfolding of the three tryptophan-containing alpha-subunits is virtually identical to the wild-type protein. These wild-type properties make these proteins attractive candidates for a fluorescence examination of the behavior of the individual folding units and the structure of potential intermediate(s) and as host proteins for the insertion of our existing destabilizing and/or stabilizing mutational alterations. SN - 0021-9258 UR - https://www.unboundmedicine.com/medline/citation/7629069/Tryptophan_containing_alpha_subunits_of_the_Escherichia_coli_tryptophan_synthase__Enzymatic_and_urea_stability_properties_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0021-9258(17)48001-X DB - PRIME DP - Unbound Medicine ER -