Dramatic Thermostabilization of Yeast Iso-1-cytochrome C by an Asparagine----isoleucine Replacement at Position 57

Overview

Journal Proc Natl Acad Sci U S A

Specialty Science

Date 1989 Jan 1

PMID 2536164

Citations 20

Authors

G Das

D R Hickey

D McLendon

G McLendon

F Sherman

Affiliations

Soon will be listed here.

Abstract

Two Saccharomyces cerevisiae yeast mutants, cyc1-73 and cyc1-190, contain nonfunctional and presumably unstable forms of iso-1-cytochrome c due to Gly-34----Ser and His-38----Pro replacements, respectively. Second-site reversions that produced Asn-57----Ile replacements at least partially restored function, presumably by alleviating the instability of these two altered iso-1-cytochromes c. Introduction of the Ile-57 replacement by site-directed mutagenesis in an otherwise normal protein resulted in a 17 degrees C increase in the transition temperature (Tm), corresponding to over a 2-fold increase in the free energy change (delta G degrees) for thermal unfolding.

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References

Ramdas L, Sherman F, Nall B . Guanidine hydrochloride induced equilibrium unfolding of mutant forms of iso-1-cytochrome c with replacement of proline-71. Biochemistry. 1986; 25(22):6952-8. DOI: 10.1021/bi00370a032. View

Russel M, Kidd S, Kelley M . An improved filamentous helper phage for generating single-stranded plasmid DNA. Gene. 1986; 45(3):333-8. DOI: 10.1016/0378-1119(86)90032-6. View

Holzschu D, Principio L, Conklin K, Hickey D, Short J, Rao R . Replacement of the invariant lysine 77 by arginine in yeast iso-1-cytochrome c results in enhanced and normal activities in vitro and in vivo. J Biol Chem. 1987; 262(15):7125-31. View

SCHELLMAN J . The thermodynamic stability of proteins. Annu Rev Biophys Biophys Chem. 1987; 16:115-37. DOI: 10.1146/annurev.bb.16.060187.000555. View

Matthews B, Nicholson H, Becktel W . Enhanced protein thermostability from site-directed mutations that decrease the entropy of unfolding. Proc Natl Acad Sci U S A. 1987; 84(19):6663-7. PMC: 299143. DOI: 10.1073/pnas.84.19.6663. View

Alber T, Sun D, Wilson K, Wozniak J, Cook S, Matthews B . Contributions of hydrogen bonds of Thr 157 to the thermodynamic stability of phage T4 lysozyme. Nature. 1987; 330(6143):41-6. DOI: 10.1038/330041a0. View

Kunkel T, Roberts J, Zakour R . Rapid and efficient site-specific mutagenesis without phenotypic selection. Methods Enzymol. 1987; 154:367-82. DOI: 10.1016/0076-6879(87)54085-x. View

Alber T, Bell J, Sun D, Nicholson H, Wozniak J, Cook S . Replacements of Pro86 in phage T4 lysozyme extend an alpha-helix but do not alter protein stability. Science. 1988; 239(4840):631-5. DOI: 10.1126/science.3277275. View

Louie G, Hutcheon W, Brayer G . Yeast iso-1-cytochrome c. A 2.8 A resolution three-dimensional structure determination. J Mol Biol. 1988; 199(2):295-314. DOI: 10.1016/0022-2836(88)90315-4. View

10.

Hampsey D, Das G, Sherman F . Yeast iso-1-cytochrome c: genetic analysis of structural requirements. FEBS Lett. 1988; 231(2):275-83. DOI: 10.1016/0014-5793(88)80834-2. View

11.

Mitchinson C, Baldwin R . The design and production of semisynthetic ribonucleases with increased thermostability by incorporation of S-peptide analogues with enhanced helical stability. Proteins. 1986; 1(1):23-33. DOI: 10.1002/prot.340010106. View

12.

Hecht M, STURTEVANT J, Sauer R . Stabilization of lambda repressor against thermal denaturation by site-directed Gly----Ala changes in alpha-helix 3. Proteins. 1986; 1(1):43-6. DOI: 10.1002/prot.340010108. View

13.

Bryan P, Rollence M, Pantoliano M, Wood J, Finzel B, Gilliland G . Proteases of enhanced stability: characterization of a thermostable variant of subtilisin. Proteins. 1986; 1(4):326-34. DOI: 10.1002/prot.340010406. View

14.

Hickey D, McLendon G, Sherman F . Thermodynamic stabilities of yeast iso-1-cytochromes c having amino acid substitutions for lysine 32. J Biol Chem. 1988; 263(34):18298-305. View

15.

Pantoliano M, Ladner R, Bryan P, Rollence M, Wood J, Poulos T . Protein engineering of subtilisin BPN': enhanced stabilization through the introduction of two cysteines to form a disulfide bond. Biochemistry. 1987; 26(8):2077-82. DOI: 10.1021/bi00382a002. View

16.

Chou P, Fasman G . Conformational parameters for amino acids in helical, beta-sheet, and random coil regions calculated from proteins. Biochemistry. 1974; 13(2):211-22. DOI: 10.1021/bi00699a001. View

17.

Chou P, Fasman G . Prediction of protein conformation. Biochemistry. 1974; 13(2):222-45. DOI: 10.1021/bi00699a002. View

18.

Sherman F, Stewart J, Jackson M, Gilmore R, Parker J . Mutants of yeast defective in iso-1-cytochrome c. Genetics. 1974; 77(2):255-84. PMC: 1213128. DOI: 10.1093/genetics/77.2.255. View

19.

Smith D, Maggio E, Kenyon G . Simple alkanethiol groups for temporary blocking of sulfhydryl groups of enzymes. Biochemistry. 1975; 14(4):766-71. DOI: 10.1021/bi00675a019. View

20.

Sherman F, Jackson M, Liebman S, Schweingruber A, Stewart J . A deletion map of cyc1 mutants and its correspondence to mutationally altered iso-1-cytochromes c of yeast. Genetics. 1975; 81(1):51-73. PMC: 1213389. DOI: 10.1093/genetics/81.1.51. View