Studies on Thiol proteinase inhibitors (cystatins) from goat liver

Abstract

The term cystatin refers to proteins that specifically inhibit the activity of newlinepapain like lysosomal cysteine proteinases. Their function is to protect the cell from unwanted proteolysis. These endogenous thiol proteinase inhibitors are widely distributed in animals, plants and microorganisms. These are non-covalent and tight binding inhibitors. Cystatins have been found to be evolutionary, structurally and functionally related forming the cystatin superfamily . On the basis of sequence homology, molecular weight, carbohydrate content and presence or absence of disulphide bonds, cystatin superfamily has been divided into three families: Family I also called as stefins include members of low molecular weight proteins (approximately 11 kDa), which lack disulphide bonds and carbohydrate content. This family includes cystatin A, B, stefin C and stefin D. Family II known as cystatins family represented by the inhibitors of a bit higher molecular weight proteins (approximately 13 kDa) as compared to stefins and possess two disulphide bonds towards carboxyl terminal. This family comprises of cystatins C, D, S, SN, E, F and M. Family III or kininogens are higher molecular weight inhibitors containing both disulphide linkages and carbohydrate content. They are found only in blood plasma. There are three distinct types of kininogens designated as high molecular weight kininogen, HK (Mr 88-120 kDa), low molecular weight kininogen LK (Mr 50-70 kDa) and T-kininogen, TK (Mr 68 kDa) which is found only in rat plasma. Cystatins have been purified from several mammalian sources like muscle, adder, hoof, brain, kidney, spleen, liver, skin, placenta and pancreas. In the present study cystatin has been purified from a very important mammalian source that is goat liver and named as liver cystatin (LC). The purification of goat liver cystatin was achieved using a simple and convenient four step procedure including alkaline treatment, acetone fractionation, ammonium sulphate precipitation and gel filtration chromatography.