The Interaction Between High- and Low-Sulphur Proteins Extracted from -Keratin

Abstract

Wool and many mammalian keratins consist of two classes of proteins, one which is higher in sulphur content than the parent keratin and is of basic character in the unmodified keratin and one which is lower in sulphur content and is acidic in character (Alexander and Earland 1950;Corfield, Robson, and Skinner 1958;Gillespie and Simmonds 1960). In the intact fibre the low-sulphur proteins are thought to occur in the microfibrils and the high-sulphur proteins in the matrix, and the two are probably linked together by disulphide bonds (Birbeck and Mercer 1957; Rogers 1959).After the disulphide bonds have been broken, by reduction followed by alkylation with iodoacetate, or by oxidation, the two classes of protein can be separated in two ways. The high-sulphur component may be preferentially extracted in the presence of salt (Corfield, Robson, and Skinner 1958;Gillespie 1962) or of certain specific ions (e.g. zinc or cetyltrimethylammonium ion) which suppress the solubility of the low-sulphur proteins (Gillespie 1962). Alternatively a mixture of the proteins can be extracted at low ionic strength and the lowsulphur proteins separated from the others by precipitation at acid pH values. It. has recently been observed that the latter separation when applied to the reduced and S-carboxymethylated proteins can be complicated by co-precipitation of the two classes of proteins. This paper presents an account of this phenomenon and a summary of conditions suitable for separating mixtures of high-and low-sulphur proteins. Results and Discussion(i) Reduced and Oarboxymethylated Proteins.-Erratic and variable yields of high-sulphur protein were obtained when the low-sulphur proteins were removed from mixtures by precipitation at pH 4·1 at an ionic strength of 0 ,1. It. has been reported that a number of reprecipitations were necessary to remove the highsulphur proteins completely from the low-sulphur protein precipitated under these conditions (Gillespie 1960). A study of artificially prepared mixtures of high-and low-sulphur proteins showed that, as the ratio of low-sulphur protein to high in the mixture increases (Fig. I), so the recovery of high-sulphur protein after precipitation falls. Treatment of the supernatant with Zn2+ at pH 6 (Gillespie 1957) showed that the low-sulphur proteins were always quantitatively precipitated.Further data on this co-precipitation of high-and low-sulphur proteins are shown in Table 1. The mixed high-and low-sulphur proteins used in this experiment were prepared from Merino wool, either by extraction with 0 ·IM potassium * Manuscript

Extraction of Reduced Wool Proteins by Solutions of Salts

“…This "standard" solution has pH 7·9 (measured with a glass electrode) and addition of alkali or acid to vary the pH in the range 5 ·0-10·2 caused no change in the extent of extraction. This extraction technique has also been applied on a preparative scale, the extracted protein being then S-carboxymethylated with iodoacetate and separated into "low-sulphur" (SCMKA) and "high-sulphur" (SCMKB) fractions by the usual fractional precipitation technique (Gillespie, O'Donnell, and Thompson 1962). In order to explore the possibility of some selective extraction of particular components from the wool fibre, preparative runs have been made using different propan-1-o1 concentrations in sodium iodide (5M) and different sodium iodide concentrations in aqueous propan-1-ol (25% vjv).…”

N-Acetyl Groups in Wool and Extracted Wool Proteins

“…In the case of the low-sulphur protein a-keratose from oxidized wool, we know that there is some contamination by a protein resembling the high-sulphur protein (cf. Thompson and O'Donnell 1962a), and the high-and low-sulphur proteins from reduced and alkylated wool can also co-precipitate (Gillespie, O'Donnell, and Thompson 1962).…”

“…In preparation C, where six separate 1-g samples of wool were reduced, four were subsequently treated as in preparations A and B while the remaining two samples were precipitated and washed with O· 4N trichloroacetic acid and then dialysed to remove residual reagents before alkylation. In preparation D the reduced wool was precipitated and washed with 0·4N trichloroacetic acid before dialysis in order to increase the percentage extraction of soluble alkylated proteins and also to recover a higher yield of high-sulphur protein (Thompson and O'Donnell 1962). For fractionation of extracted 8-carboxymethyl proteins into high-(SCMKB) and low-sulphur (SCMKA) fractions two methods were used.…”

Studies on Oxidized Wool VI. Interactions Between High- and Low-'Sulphur Proteins and their Significance in the Purification of Extracted Wool Proteins

“…The data in Table 4 suggest that the percentage of high-sulphur protein in wool may be of the order of 40% by weight if it is remembered that about 20% of the wool remains undissolved and could contain some high-sulphur protein. The preferential extraction of highsulphur proteins from reduced wool has to date only been of the order of 25% by weight of the original wool (Gillespie 1962).…”