1. A method is described for the preparation from Actinidia chinensis (Chinese gooseberry) fruit of a crystalline proteinase which catalyzes the hydrolysis of N2-benzoyl-L-arginine ethyl ester, N2-p-toluene-sulphonyl-L-glutamine p-nitrophenyl ester (used in the routine assay) and a t least 15 o f the peptide bonds of gelatin. The synthesis of the tosyl-glutamine p-nitrophenyl ester is described. The enzyme resembles papain in its action against benzoyl-L-arginine ethyl ester, having a broad pH-optimum from pH 5-7, with K,(app) of 89 mM and keat of about 2.6 sec-l at pH 5.6, 25" in 0.3 M KCI.2. The enzymic activity elutes from Sephadex G-50 as a protein of molecular weight 12800 f 700, and is partially resolved from inactive protein of apparent molecular weight 15400 f 800. The preparation has minimum solubility near pH 3.1 and migrates a t both pH 8.3 and 5.0 on polyacrylamide gel electrophoresis as two closely-spaced anionic bands. These two electrophoretic components are partially resolved by chromatography on DEAE-cellulose and both are enzymically active against tosyl-L-glutamine p-nitrophenyl ester.3. The inhibitions of enzymic activity by 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB) and 4,4'-dithiodipyridine, both reversible by 1,4-dithiothreitol, show that enzymic activity is dependent on an intact sulphhydryl group assaying about 0.23 moles per 12800 g protein. The enzyme is rapidly inactivated by iodoacetate and thereafter gives a low sulphhydryl assay with dithiodipyridine. Conversely, pretreatment with DTNB gives substantial protection against iodoacetate.The proteinase activity in extracts of New Zealand commercial strains of the ripe fruit of the Chinese Gooseberry, Actinidia chinensis has been characterized by Arcus [1] by its activity against gelatin (optimum p H 4 ) and haemoglobin, and was shown to be enhanced by cysteine, Na,S, KCN and EDTA. It is of interest to isolate the enzyme(s) responsible for this activity because of possible structural homologies with the sulphhydryl plant proteinases papain, ficin, bromelin, chymopapain B and pinguanain [2,3], especially now that the tertiary structure of crystalline papain has been elucidated [4].This report describes the preparation of a crystalline anionic proteinase from A. chinensis, some physical and enzymic properties, and further evidence for the requirement of a sulphhydryl group for activity.Unusual Abbreviations. DTNB, 5,5'-dithiobis-(2-nitrobenzoic acid); CNps-proteinase, 3-carboxy-4-nitrophenylsulphenyl-proteinase; BzArgOEt, N2-benzoyl-L-arginine ethyl ester hydrochloride ; TosGlnONp, N2-p-toluenesulphonyl-L-glutamine p-nitrophenyl ester; Pr,P-trypsin, diisopropylphosphoryl-trypsin ; V , and V,, void volume and elution volume.

JUNE 1963 .oj~l.l I 465 set. Thus, except in the case of the T -set condition, the fibers were exposed to water at 97°C for 4 hr, and therefore the differences in fiber structure of the p-set and P-T-set fibers must be due to the mechanical restraint conditions imposed during the treatments. The fact that the P-T fibers are stronger and less extensible than the corresponding P-set fibers suggests a more perfect network of secondary and primary bonding in the P-T condition. It would also suggest that relaxation at the temperature of boiling water causes some breakdown of network bonding with subsequent deorientation.The differences in behavior among the four keratin fiber types are relatively minor, although in many instances si~ificant and i~triguing.~hese differences in behavior may possibly be associated ith the varied cortical structures, although definite ;:Isignments in regard to the effect of setting on the h anical properties of the ortho-and the paramec h basi f cortex are not possible on t e asis 0 these results. AbstractFour types of wool and three types of mohair were examined by differential thermal analysis (DTA) in the natural and dried states. The DTA curves of wool and mohair were not significantly different. Curves for both in the natural state showed an endotherm at 130-145°, ascribed to the vaporization of bound water, and endotherms at 220-230°and 230-250°accompanied by charring, the release of gaseous decomposition products, and, ultimately, liquefaction. Derivatives of one type of wool and one type of mohair prepared by acetylation or modification of the disulfide cross-linkages were compared with unmodified materials by DTA. Differences in the DTA curves are described, and possible correlations of these with the changes in chemical structure are suggested. Collagen, gelatin, silk, horn keratin, cystine, and cysteine hydrochloride were also characterized. AbstractA. method. for ca~rying out potenttometric~itration of reduced vat dyes in aqueous alkaline solutions usmg a potassium ferricyanide solution as the oxidant is described The standard method is modified on the basis of certain assumptions, and the values of potentials~re~lculated and termed.".01aracteristic redox potentials." An account of the deter':lIn~tlOn of these charac~e\lslc redox potential values for a series of purified vat dyes IS given. The.characteristie redox potential values of vat dyes are compared with the ease of reduction of these dyes and also with their tendering behavior. The effect of substitution on these values is discussed.

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