title: Influences of amino acid features of glutathione S-transferase fusion proteins on their solubility.
abstract: We have previously described our strategy for high-throughput (HT) production of recombinant antigens for anti-mKIAA antibody generation, which involves using shotgun fragments generated during entire sequencing of mKIAA cDNAs. We applied this strategy to 1628 mouse KIAA (mKIAA) cDNA fragments, and 84.2% of the GST-mKIAA fusion proteins were successfully purified. The solubility of the proteins was predicted by a small-scale bacterial culture, and a large-scale culture was then performed according to the expected results. Among them, 43.8% of the proteins were purified as a soluble form and 56.2% as an insoluble form. The average yield of the soluble proteins was 0.15 nmol/mL of bacterial culture, and that of the insoluble proteins was 0.55 nmol/mL Statistical analysis of the data revealed a significant correlation between amino acid features of the recombinant proteins and their solubility. To achieve the most effective and feasible protein expression, we constructed a decision tree in which the analyzed data were reflected. The information described here may provide practical guidelines for HT production of recombinant proteins.
authors.affiliation: Chiba Industry Advancement Center, Mihama-ku, Chiba, Japan.
authors.forename: Kiyo
authors.initials: K
authors.lastname: Shimada
authors.forename: Mihoko
authors.initials: M
authors.lastname: Nagano
authors.forename: Makoto
authors.initials: M
authors.lastname: Kawai
authors.forename: Hisashi
authors.initials: H
authors.lastname: Koga
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry*; D000818:Animals; D000941:Antigens; D018076:DNA, Complementary / Q000235:genetics; D003663:Decision Trees; D005982:Glutathione Transferase; D006801:Humans; D051379:Mice; D008970:Molecular Weight; D011993:Recombinant Fusion Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D012995:Solubility
chemical_list: D000596:Amino Acids; D000941:Antigens; D018076:DNA, Complementary; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase