title: Efficient production of native, biologically active human cystatin C by Escherichia coli.
abstract: A cDNA encoding the mature human cysteine proteinase inhibitor cystatin C was fused to the coding sequence for the Escherichia coli outer membrane protein A signal peptide, and the recombinant gene was expressed in E. coli under the control of the lambda PR promoter, an optimized Shine-Dalgarno sequence and the lambda cI 857 repressor. When induced at 42 degrees C, such cells expressed large amounts of recombinant cystatin C. The recombinant protein was isolated in high yield and characterized. All physicochemical properties investigated, including the positions of disulfide bonds, indicated that the E. coli derived cystatin C was identical to cystatin C isolated from human biological fluids, except that the proline residue in position three was not hydroxylated. The recombinant protein displayed full biological activity against papain, cathepsin B and dipeptidyl peptidase I.
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D055316:Cystatin C; D015891:Cystatins; D004247:DNA / Q000235:genetics; D004587:Electrophoresis, Agar Gel; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D008969:Molecular Sequence Data; D011480:Protease Inhibitors / Q000096:biosynthesis* / Q000235:genetics; D014176:Protein Biosynthesis; D021382:Protein Sorting Signals / Q000235:genetics; D011506:Proteins / Q000235:genetics; D011507:Proteinuria; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics
chemical_list: C525030:CST3 protein, human; D055316:Cystatin C; D015891:Cystatins; D011480:Protease Inhibitors; D021382:Protein Sorting Signals; D011506:Proteins; D011994:Recombinant Proteins; D004247:DNA