Isolation and characterization of a unique protease from sporulating cells of Bacillus subtilis

Two proteases, designated I and II, have been isolated from sporulating cells of Bacillus subtilis. They were partially purified by ammonium sulfate fractionation, Sephadex chromatography and affinity columns. Protease I was found to be similar to an already characterized B. subtilis protease. Protease II is trypsin-like in its substrate specificity and is distinct from protease I in its pH optimum, pH stability, molecular weight, substrate specificity, heat stability and sensitivity to various inhibitors. While both enzymes were produced primarily during sporulation, they attained maximum levels of activity at different times. Distinct functions for these proteases in post exponential B. subtilis are likely.     

A study of the pollen grains of Amaranthus spinosus Linné and A. Dubius Mart ex Thellung and their hybrids

A pollen-morphology study of Amaranthus spinosus, A. dubius, and their hybrids has been carried out. Three pollen types have been observed, namely (1) Type A: micrograins; (2) Type B: grains with smaller pores; and (3) Type C: grains with larger pores. Type B is characteristic of A. spinosus, Type C of A. dubius, and the micrograins of the hybrids. Pollen size range, and frequency of the various morphotypes serve to throw light on the biosystematics of the plants studied.     

Modification of the rapidly reacting thiols of atrial and ventricular myosins. Evidence for structural variances around the SH-1 thiol of the two isozymes

Environmental variances were noted around the rapidly reacting SH-1 thiol of ventricular versus atrial myosin, based on electron paramagnetic resonance studies. Further studies, in which either SH-1 or SH-1 + SH-2 thiols were modified with N-ethylmaleimide, indicated the importance of the SH-2 moiety of both isozymes for generation of tension, when analyzed as synthetic actomyosin threads. Comparative EPR studies showed that the spin label was more strongly immobilized when complexed to ventricular SH-1 thiol as compared to when it was complexed to atrial myosin. Likewise, addition of PPi, ATP or ADP created a greater mobility in the spin label when added to ventricular spin-labeled myosin as compared to that of atrial myosin. comparative studies of spin-labeled actomyosin versus myosin analyzed at different EPR power settings also demonstrated disparaties surrounding the SH-1 thiol between the two myosin isozymes.     

Extractive lactic acid fermentation using ion-exchange resin

Lactic acid fermentation is an end-product-inhibited reaction. The restriction imposed by lactic acid on its fermentation can be avoided by extractive fermentation techniques. Studies were performed by attaching an ion-exchange resin packed column with a 2-L fermentor for separation of lactic acid. The fermentation, in a conventional batch mode, resulted in a lactic acid yield of 0.828 g . g(-1) and a lactic acid productivity of 0.313 g . L(-1) . h(-1). However, these could be further enhanced to 0.929 g . g(-1) and 1.665 g . L(-1) . h(-1) by extractive fermentation techniques. The effect of temperature on extractive fermentation was remarkable and has been included in this work.     

The fine structure of psychrophilic and mesophilic Rhodotorula rubra

Transmission electron microscopy of thin sections of two mesophilic and a psychrophilic Rhodotorula rubra revealed structures normally associated with yeast. The capsule of all three strains was thicker in the cells grown in glucose than those grown in glucose-free media. The cell wall of all three strains showed more lamellae in the cells grown in a medium containing glycerol. Budding was of the Rhodoturula type. Endoplasmic reticula running parallel to the nucleus were commonly observed in the psychrophiles but not in the mesophiles. In the psychrophile, in association with the plasmalemma, lomasomes and plasmalemmasomes were observed. Giant mitochondria were commonly seen in the cells grown in a fermentable carbohydrate-free medium. Vacuoles were mostly empty.     

A note on suxamethonium sensitivity and serum cholinesterase variants

Summary Sera from 21 cases of prolonged apnoea which showed normal phenotype (UU) on the basis of dibucaine and fluoride inhibition were re-examined by replacing the substrate benzoylcholine with succinylcholine (suxamethonium). 9 samples had normal enzyme activity but low dibucaine number (DN=<20) indicating the atypical variant; 6 sera showed no detectable enzyme activity. The remaining 6 samples had enzyme activity and DN comparable with healthy controls. The occurence of new variants of serum cholinesterase sensitive only to succinylcholine is suggested.Dedicated to Prof. Dr. med. J. Kühnau on his 75th birthday.     

Purification of bull sperm hyaluronidase by concanavalin-A affinity chromatography.

A new method for obtaining highly purified hyaluronidase (hyaluronate glycanohydrolase EC 3.2.1.25) in high yield is described. Bull seminal plasma was fractionated with (NH4)2 SO4 and the 30 to 65% saturation fractions were applied to a DEAE-cellulose column. The first protein peak contained hyaluronidase, beta-N-acetylglucosaminidase and beta-glucuronidase. The latter two enzymes were separated by gel filtration on Sephadex G-200. The hyaluronidase was further purified by a Concanavalin-A Sepharose 4B affinity column. By gradient elution with alpha-methyl-D-glucoside a fraction which had a specific activity of 1998 units/mg protein (57 942 National Formulary Standard units/mg protein) was obtained. The highly purified enzyme showed one major protein band on acrylamide gel electrophoresis at pH 4.3. The purified hyaluronidase did not show any beta-glucuronidase or beta-N-acetylglucosaminidase activities. The percent yield of purified hyaluronidase calculated on the basis of total activity was ten times higher than by any pervious method [Yang, C.H. and Srivastava, P.N. (1975) J. Biol. Chem. 250, 79-83].     

Inhibition of bull and rabbit sperm enzymes by alpha-chlorohydrin.

High concentrations of alpha-chlorohydrin were found to inhibit hyaluronidase, beta-glucuronidase, and aryl sulphatases in bull and rabbit spermatozoa, but not acrosin and neuraminidase. Preincubation of the enzyme and alpha-chlorohydrin was essential to achieve the maximum inhibition which was irreversible.