SummaryThe oxidation of wool with performic and peracetic acids has been compared by measuring the cysteic acid contents of hydrolysates of the oxidized wool. Whereas virtually complete oxidation takes place with performic acid, the oxidation with peracetic acid is incomplete. Performic acid can also further oxidize the partial oxidation products in wool treated with peracetic acid and it is concluded that performic acid is the more powerful oxidizing agent for wool. For the oxidation of the disulphide bonds prior to the extraction of proteins from wool, performic acid is therefore the better reagent.

The amino acid sequence of the major components of the silver gull feather calamus has been determined and compared with that of the emu. The sequenator was used with a modified Edman-Begg program to facilitate determination of the sequence of the large hydrophobic fragment obtained on tryptic digestion. The main features of the comparison were: (1) the overall structures of the polypeptide chains were similar, having non-crystalline cystine-rich sections towards either end of the chain separated by a large crystalline region of 62 residues which contained the majority of the hydrophobic and serine and glycine residues; (2) approximately one-sixth of the residues were different in the two species, with the majority of changes occurring in the tails (i.e. non-crystalline or matrix region). The data argue for stringent demands in the selection of amino acids for the crystalline part of the feather molecule, a severity that is probably comparable to the strict requirements for the sequence of some of the enzymes. 1 1 1 S 0·2 J J "

SummaryN . acetyl groups have been shown to exist in wool and in proteins extracted from wool after reduction with mercaptoethanol followed by alkylation with iodoacetate or after oxidation with performic acid. The evidence suggests that these acetyl groups are located on N-terminal amino groups of peptide chains rather than on the E-amino groups of lysine.Acid hydrolysis with 12N sulphuric acid was found to be preferable to alkaline hydrolysis for the release of acetic acid from the N-acetyl groups, since alkaline hydrolysis produced volatile acids other than acetic acid.

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

SummartJStarch-gel electrophoresis in buffers containing 8ror urea has been used to follow the fractionation of wool proteins extracted from reduced and carboxymethylated wool. Fractionation on both DEAE-cellulose and Sephadex G-200 in the presence of buffers containing Sr.t: urea is possible but in neither case is a single component obtained. However, a combination of these two methods has enabled one of the major components to be isolated. The amino acid composition of this material is reported.

Two amino terminal acetyl peptides, N�acetyl Ser.Phe.Asp.Phe (A) and N�acetyl Ser.Tyr.Asp.Phe (B), have been isolated from component 8, one of the two major components present in extracts of reduced and carboxymethylated wool. These peptides could not be eluted from columns of Dowex 1 but could be satis� factorily fractionated by DEAE-cellulose chromatography with a gradient of formic acid and by paper ionophoresis. The yield of peptide A was approximately twice that of peptide B. If the molecular weight of component 8 is 45,000 then these acetyl peptide sequences account for only approximately half of the theoretical amount of terminal peptides. This low recovery and the presence of two different but related terminal peptides provides additional evidence for heterogeneity of component 8, and support the idea .that a family of related proteins is present in keratin.

The variables involved in the alkaline extraction of wool oxidized with performic acid have been studied and reproducible methods of extraction have been developed. The presence of potassium sulphate reduces the amount of protein extracted but, for maximal precipitation of protein at pH 4, potassium sulphate is necessary. The extraction has been followed by estimating the cysteic acid and nitrogen contents of the soluble material. It is shown that the fractions of oxidized wool-a- +y-, a-, and y-keratose-contain approximately 9, 5, and 16 g cysteic acid nitrogen per 100 g total nitrogen respectively, whether extraction is carried out at pH 8 or II.

SummaryStarch-gel electrophoresis of wool proteins extracted from reduced and carboxymethylated wool gives a complex pattern in which there are two major protein bands. By a combination of chromatography on DEAE-cellulose and gelfiltration in buffers containing 8M urea these two protein components have been isolated. The amino acid composition and some properties of these two fractions are reported. A comparison of the amino acid composition and of peptide maps of tryptic digests of the two fractions shows distinct differences between them, and by labelling with 2-[14C]iodoacetate the distribution of the peptides containing S-carboxymethylcysteine residues were also shown to be different.The molecular weight of the two protein components was estimated to be about 45,000 by comparison of their elution volumes on gel-filtration with the elution volumes of reduced and carboxymethylated proteins of known molecular weight. Gel-filtration was carried out in 8M urea, 14M formamide, and 5M guanidine hydrochloride, and no evidence of further dissociation of these components was obtained.