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Purification and Characterization of a Chicken Egg White Cystatin Variant Expressed in an Escherichia Coli pIN-III-ompA System

Purification and Characterization of a Chicken Egg White Cystatin Variant Expressed in an Escherichia Coli pIN-III-ompA System

E A Auerswald, G Genenger, R Mentele, S Lenzen, I Assfalg-Machleidt, L Mitschang, H Oschkinat, H Fritz

Eur J Biochem. 1991 Aug 15;200(1):131-8.

PMID: 1879418

Abstract:

A synthetic gene coding for a chicken egg white cystatin variant was cloned and expressed using the pIN-III-ompA Escherichia coli expression system. After osmotic shock of the E. coli cells, the cysteine proteinase inhibitor was isolated from periplasm and purified by S-carboxymethylpapain affinity chromatography. The resulting inhibitory material was characterized by SDS/PAGE, reversed-phase HPLC, peptide mapping and amino acid sequencing. The recombinant variant chicken AEF-[S1----M, M29----I, M89----L]cystatin shows strong inhibitory activity and displays Ki values in the complex with papain, actinidin and cathepsin B similar to those found for natural chicken cystatin. The purified variant showed a native-chicken-cystatin-like conformational state, as determined by NMR spectroscopy, if the NMR data of 15N-labelled recombinant inhibitor were compared with those of the natural inhibitor.

Chemicals Related in the Paper:

Catalog Number	Product Name	Structure	CAS Number	Price
AP81989959	Cystatin from chicken egg white		81989-95-9	Price

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