title: Application of SGT1-Hsp90 chaperone complex for soluble expression of NOD1 LRR domain in E. coli.
abstract: NOD1 is an intracellular sensor of innate immunity which is related to a number of inflammatory diseases. NOD1 is known to be difficult to express and purify for structural and biochemical studies. Based on the fact that Hsp90 and its cochaperone SGT1 are necessary for the stabilization and activation of NOD1 in mammals, SGT1 was chosen as a fusion partner of the leucine-rich repeat (LRR) domain of NOD1 for its soluble expression in Escherichia coli. Fusion of human SGT1 (hSGT1) to NOD1 LRR significantly enhanced the solubility, and the fusion protein was stabilized by coexpression of mouse Hsp90α. The expression level of hSGT1-NOD1 LRR was further enhanced by supplementation of rare codon tRNAs and exchange of antibiotic marker genes.
authors: School of Chemical and Biological Engineering, Seoul National University, 1 Gwanak-ro, Gwanak-gu, Seoul 08826, Republic of Korea.
authors: Tae-Joon
authors: TJ
authors: Hong
authors: School of Chemical and Biological Engineering, Seoul National University, 1 Gwanak-ro, Gwanak-gu, Seoul 08826, Republic of Korea. Electronic address: hahnjs@snu.ac.kr.
authors: Ji-Sook
authors: JS
authors: Hahn
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D018841:HSP90 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D015151:Immunoblotting; D051379:Mice; D053474:Nod1 Signaling Adaptor Protein / Q000235:genetics / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D055550:Protein Stability; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D020449:Repetitive Sequences, Amino Acid / Q000235:genetics; D012995:Solubility
chemical_list: D018797:Cell Cycle Proteins; D018841:HSP90 Heat-Shock Proteins; C107835:Hspca protein, mouse; C119067:NOD1 protein, human; D053474:Nod1 Signaling Adaptor Protein; D011993:Recombinant Fusion Proteins; C490164:SUGT1 protein, human
keywords: Heat shock protein 90 kDa (Hsp90); Leucine-rich repeat (LRR); Nucleotide-binding oligomerization domain protein 1 (NOD1); Soluble expression; Suppressor of G2 allele of skp1 (SGT1)