Expression, purification, and initial structural characterization of YadQ, a bacterial homolog of mammalian ClC chloride channel proteins.
YadQ of Escherichia coli is a homolog of the mammalian chloride channels of the ClC family. The yadQ gene was cloned as a fusion protein with a hexahistidine tag and tobacco etch virus protease site for the removal of the tag. The protein was expressed in the membrane of E. coli and extracted with decylmaltoside. Purification was achieved by metal affinity chromatography followed by cation exchange. Circular dichroism revealed a high alpha-helical content. Size exclusion chromatography suggests that YadQ forms dimers. The similarity in primary, secondary, and quaternary structure and the ability to recombinantly express YadQ in the cell membrane make the protein a good candidate for the structural study of ClC chloride channels.
Purdy
M D
MD
Biophysics Program, University of Virginia, 1300 Jefferson Park Ave, Charlottesville, VA 22906-0011, USA.
Wiener
M C
MC
D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification*; D001483:Base Sequence; D018118:Chloride Channels / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification*; D002942:Circular Dichroism; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D019281:Dimerization; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D005798:Genes, Bacterial; D006801:Humans; D008969:Molecular Sequence Data; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D017386:Sequence Homology, Amino Acid
D001426:Bacterial Proteins; D018118:Chloride Channels; D017931:DNA Primers; D011993:Recombinant Fusion Proteins