The CXC motif: a functional mimic of protein disulfide isomerase.
Protein disulfide isomerase (PDI) utilizes the active site sequence Cys-Gly-His-Cys (CGHC; E degrees ' = -180 mV) to effect thiol-disulfide interchange during oxidative protein folding. Here, the Cys-Gly-Cys-NH(2) (CGC) peptide is shown to have a disulfide reduction potential (E degrees ' = -167 mV) that is close to that of PDI. This peptide has a thiol acid dissociation constant (pK(a) = 8.7) that is lower than that of glutathione. These attributes endow the CGC peptide with substantial disulfide isomerization activity. Escherichia coli thioredoxin (Trx) utilizes the active site sequence Cys-Gly-Pro-Cys (CGPC; E degrees ' = -270 mV) to effect disulfide reduction. Removal of the proline residue from the Trx active site yields a CGC active site with a greatly destabilized disulfide bond (E degrees ' >or= -200 mV). The DeltaP34 variant retains high conformational stability and remains a substrate for thioredoxin reductase. In contrast to the reduced form of the wild-type enzyme, the reduced form of DeltaP34 Trx has disulfide isomerization activity, which is 25-fold greater than that of the CGC peptide. Thus, the rational deletion of an active site residue can bestow a new and desirable function upon an enzyme. Moreover, a CXC motif, in both a peptide and a protein, provides functional mimicry of PDI.
Department of Biochemistry, University of Wisconsin-Madison, Madison, Wisconsin 53706, USA.
Kenneth J
KJ
Woycechowsky
Ronald T
RT
Raines
D020816:Amino Acid Motifs; D001665:Binding Sites; D003545:Cysteine / Q000737:chemistry; D004220:Disulfides / Q000737:chemistry / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology* / Q000378:metabolism*; D005822:Genetic Vectors; D007700:Kinetics; D018716:Molecular Mimicry; D009154:Mutation; D009249:NADP / Q000378:metabolism; D010084:Oxidation-Reduction; D011487:Protein Conformation; D019704:Protein Disulfide-Isomerases / Q000737:chemistry / Q000378:metabolism*; D017510:Protein Folding; D013050:Spectrometry, Fluorescence; D013379:Substrate Specificity; D013438:Sulfhydryl Compounds / Q000737:chemistry / Q000378:metabolism; D013880:Thioredoxin-Disulfide Reductase / Q000378:metabolism; D013879:Thioredoxins / Q000235:genetics / Q000378:metabolism*
D004220:Disulfides; D013438:Sulfhydryl Compounds; D013879:Thioredoxins; D009249:NADP; D013880:Thioredoxin-Disulfide Reductase; D019704:Protein Disulfide-Isomerases; D003545:Cysteine