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Analysis of the Yeast De Novo Protein YNR034W‐A
Author(s) -
Shi Aishan,
Cordes Matthew
Publication year - 2013
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.27.1_supplement.996.15
Subject(s) - gene , biology , protein folding , frameshift mutation , yeast , genetics , biochemistry , mutation
The general model for gene evolution is that new genes are derived from old genes by speciation or duplication. Recently however, a large number of de novo genes have been discovered. YNR034W‐A is a yeast gene that has two alleles. The long sequence is partial de novo protein that is 24 residues longer than the ancestral protein (98 residues) with a different sequence after residue 65. This project aims to understand the effects of the frameshift mutation in the ancestral gene on protein folding of the partial de novo protein. Both alleles were codon‐optimized, sub‐cloned into N‐terminal polyHis‐tagged vectors, and over‐expressed in E. coli. Protein was purified by lysis, nickel affinity chromatography, and dialysis into refolding buffer. Far UV circular dichroism determined secondary structure and thermal melts were used to determine thermal stability. The longer protein is less stable in solution and more aggregation prone due to the C‐terminal tail. Further studies include truncation of the long protein at positions 65 and 98. The data support the hypothesis that de novo proteins are intrinsically disordered; these proteins provide a useful platform for understanding protein folding and evolution.