Evolution of an Arbitrary Sequence in Solubility

Overview

Journal J Mol Evol

Specialty Biochemistry

Date 2004 Mar 26

PMID 15042340

Citations 14

Authors

Yoichiro Ito

Toshihiro Kawama

Itaru Urabe

Tetsuya Yomo

Affiliations

Soon will be listed here.

Abstract

We have investigated the evolvability of an insoluble random polypeptide, RP3-34, to a soluble form through iterative mutation and selection with the aid of the green fluorescent protein (GFP) folding reporter. To assess the solubility of the polypeptides in the selected clones of each generation, the polypeptide genes were detached from the GFP fusions and expressed with a His(6) tag. The solubility of the variant random polypeptides increased in each generation within the scope of the evolutionary process, and the polypeptides assumed a soluble form from the fourth generation. Analysis of the synonymous and nonsynonymous mutations found in the deduced amino acid sequence of the selected polypeptides revealed that selection had accelerated the evolutionary rate. The solubility and hydrophobicity of the polypeptides and the 25 arbitrarily chosen random polypeptides found in a previously prepared library were determined, analyzed, and interpreted from the landscape on the protein sequence space. This study showed the evolvability of an insoluble arbitrary sequence toward a soluble one, hence, it provides a new perspective on the field of artificial evolution.

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References

Wilkinson D, HARRISON R . Predicting the solubility of recombinant proteins in Escherichia coli. Biotechnology (N Y). 1991; 9(5):443-8. DOI: 10.1038/nbt0591-443. View

KYTE J, Doolittle R . A simple method for displaying the hydropathic character of a protein. J Mol Biol. 1982; 157(1):105-32. DOI: 10.1016/0022-2836(82)90515-0. View

Maxwell K, Mittermaier A, Forman-Kay J, Davidson A . A simple in vivo assay for increased protein solubility. Protein Sci. 1999; 8(9):1908-11. PMC: 2144404. DOI: 10.1110/ps.8.9.1908. View

Hayashi Y, Sakata H, Makino Y, Urabe I, Yomo T . Can an arbitrary sequence evolve towards acquiring a biological function?. J Mol Evol. 2003; 56(2):162-8. DOI: 10.1007/s00239-002-2389-y. View

Yamauchi A, Yomo T, Tanaka F, Prijambada I, Ohhashi S, Yamamoto K . Characterization of soluble artificial proteins with random sequences. FEBS Lett. 1998; 421(2):147-51. DOI: 10.1016/s0014-5793(97)01552-4. View

Yamauchi A, Nakashima T, Tokuriki N, Hosokawa M, Nogami H, Arioka S . Evolvability of random polypeptides through functional selection within a small library. Protein Eng. 2002; 15(7):619-26. DOI: 10.1093/protein/15.7.619. View

Waldo G, Standish B, Berendzen J, Terwilliger T . Rapid protein-folding assay using green fluorescent protein. Nat Biotechnol. 1999; 17(7):691-5. DOI: 10.1038/10904. View

Wigley W, Stidham R, Smith N, HUNT J, Thomas P . Protein solubility and folding monitored in vivo by structural complementation of a genetic marker protein. Nat Biotechnol. 2001; 19(2):131-6. DOI: 10.1038/84389. View

Ito Y, Suzuki M, Husimi Y . A novel mutant of green fluorescent protein with enhanced sensitivity for microanalysis at 488 nm excitation. Biochem Biophys Res Commun. 1999; 264(2):556-60. DOI: 10.1006/bbrc.1999.1541. View

10.

Yanagihara N, Suwa M, Mitaku S . A theoretical method for distinguishing between soluble and membrane proteins. Biophys Chem. 1989; 34(1):69-77. DOI: 10.1016/0301-4622(89)80043-2. View

11.

Prijambada I, Yomo T, Tanaka F, Kawama T, Yamamoto K, Hasegawa A . Solubility of artificial proteins with random sequences. FEBS Lett. 1996; 382(1-2):21-5. DOI: 10.1016/0014-5793(96)00123-8. View

12.

Aita T, Husimi Y . Adaptive walks by the fittest among finite random mutants on a Mt. Fuji-type fitness landscape. II. Effect of small non-additivity. J Math Biol. 2000; 41(3):207-31. DOI: 10.1007/s002850000046. View

13.

Miki T, Yasukochi T, Nagatani H, Furuno M, Orita T, Yamada H . Construction of a plasmid vector for the regulatable high level expression of eukaryotic genes in Escherichia coli: an application to overproduction of chicken lysozyme. Protein Eng. 1987; 1(4):327-32. DOI: 10.1093/protein/1.4.327. View

14.

Arakawa T, Jongsareejit B, Tatsumi Y, Tanaka K, Ikeda K, Komatsubara H . Application of N-terminally truncated DNA polymerase from Thermus thermophilus (delta Tth polymerase) to DNA sequencing and polymerase chain reactions: comparative study of delta Tth and wild-type Tth polymerases. DNA Res. 1996; 3(2):87-92. DOI: 10.1093/dnares/3.2.87. View