Topographical studies on intestinal microvillous leucine β-naphthylamidase on the outer membrane surface

Summary The location of leucine -naphthylamidase on the outer surface of the microvillous membrane of rabbit small intestine was examined by analyzing the interaction of antibodies against leucine -naphthylamidase or another microvillous enzyme, sucrase-isomaltase complex, with microvillous vesicles having different relative amounts of these enzymes, in respect to vesicle agglutination, inhibition of enzyme activity, and electron-microscopic morphology. The results obtained indicate that leucine -naphthylamidase, or at least its antigenic sites, protrude about 10 nm from the outer surface of the microvillous membrane.     

Synthesis of a polymide containing a glucose unit in the main chain

A new type polyamide containing a glucose unit in the main chain has been synthesized by the polymerization of C₁, C₃, C₄ blocked C₆-carboxymethylglucosamine, prepared from chitin. The deblocking procedure gave the water-soluble polyamide, of MW 1.5 × 10⁴, which can be regarded as a model for the recognition site of lectin.     

Actin polymerization promoted by a heptapeptide, an analog of the actin-binding S site on myosin head

Polymerization of G-actin to F-actin was indicated by an increase in light-scattering intensity after the addition of a heptapeptide (Ile-Arg-Ile-Cys(MT)-Arg-Lys-Gly-OEt), an analog of the actin-binding S-site on S-1 heavy chain. The half-maximal concentration of the heptapeptide which induced an increase in the light-scattering intensity at 25 degrees C was about 110 microM, which was in the range of the dissociation constant of this peptide with F-actin. The polymerization of G-actin to F-actin by binding of the heptapeptide was further demonstrated by ultracentrifugal separation, Pi liberation, and electron microscopy. The polymerization of G-actin was induced only by the heptapeptide, but not by fragments of the heptapeptide. The well known acceleration of polymerization of G-actin by the myosin head may be due to the binding of G-actin with the S-site on the myosin head.     

Ca2(+)-calmodulin-dependent protein kinase II phosphorylates various types of non-epithelial intermediate filament proteins

We have investigated the actions of Ca2(+)-calmodulin (CaM)-dependent protein kinase II on various types of non-epithelial intermediate filament proteins, vimentin, desmin, glial fibrillary acidic protein (GFAP) and neurofilament triplet proteins. Most of these filament proteins could serve as substrates. The effects of phosphorylation on the filamentous structure of vimentin were investigated in sedimentation experiments and by using electron microscopy. The amount of unassembled vimentin increased linearly with increased phosphorylation. However, the extent of the effect of phosphorylation on the potential to polymerize was also affected by the MgCl2 concentration, under conditions for reassembly. The actions of Ca2(+)-CaM-dependent protein kinase II on non-epithelial intermediate filaments under physiological conditions are given attention.     

Temperature dependency of induction of sexual agglutinability by α pheromone in the yeast Saccharomyces cerevisiae

When pheromone-pretreated cells of an inducible a strain of Saccharomyces cerevisiae carrying the inducible gene saa1 were incubated in a growth medium at 28°C, induction of sexual agglutinability began after a 10 min lag period. If the cells were incubated at 38°C during the lag period, no induction occurred even after incubation at 28°C. Contrary to this, if the cells were incubated at 28°C during the lag period, almost complete induction occurred, even after transfer to 38°C. Temperature shift experiments revealed that 5 min incubation at 28°C was necessary for the initiation of the temperature-sensitive period and further 5 min incubation for the completion of the period. The temperature-sensitive period was sensitive to phenylmethylsulfonyl fluoride.Non-common abbreviations PBS 10^-2 M phosphate buffer solution, pH 5.5 - PMSF phenylmethylsulfonyl fluoride     

pKJ1, a naturally occurring conjugative plasmid coding for toluene degradation and resistance to streptomycin and sulfonamides.

Pseudomonas sp. TA8 isolated by m-toluate enrichment from an aqueous sample metabolized toluene and m- and p-xylenes via the meta cleavage pathway, and manifested specific resistance to streptomycin and sulfonamides. A variety of experiments revealed that the pKJ1 plasmid of about 150 megadaltons carried by TA8 specified both the toluene and xylene degradative function (the Tol function) and streptomycin/sulfonamide resistance. The deletion of a segment of pKJ1 (about 22 megadaltons) resulted in the loss of the Tol function. pKJ1 was not assigned to Pseudomonas incompatibility group P-1, P-2, P-3, or P-9.     

Disintegration of Rhodospirillum rubrum chromatophore membrane into photoreaction units, reaction centers, and ubiquinone-10 protein with mixture of cholate and deoxycholate.

1. The membrane of Rhodospirillum rubrum chromatophores was disintegrated with mild detergents (cholate and deoxycholate) in order to study the spatial arrangement of the functional proteins in the photochemical apparatus and the electron transport system in the membrane. 2. The components solubilized from the membrane by a mixture of cholate and deoxycholate (C-DOC) were separated into four fractions by molecular-sieve chromatography in the presence of C-DOC; they were designated as F1, F2, F3, and F4 in the order of elution. The fractions were further purified by repeated molecular-sieve chromatography in the presence of C-DOC until each fraction was chromatographically homogeneous. 3. F1 appeared to be conjugated forms of F2. 4. The purified F2 was composed of a rigid complex having a weight of 7 X 10(5) daltons, containing approximately 10 different kinds of protein species with molecular weights of 3.8 X 10(4), 3.6 X 10(4), 3.5 X 10(4), 2.8 X 10(4), 2.7 X 10(4), 2.6 X 10(4), 1.3 X 10(4), 1.2 X 10(4), 1.1 X 10(4), and 1.0 X 10(4). The complex contained 33 bacteriochlorophylls, 4 iron atoms, and 90 phosphates, but no cytochrome, ubiquinone, or phospholipid. It showed the same reaction center activity as chromatophores, indicating that the complex was a unit of the photochemical apparatus (photoreaction unit). Each chromatophore of average size was estimated to possess about 24 photoreaction units. 5. The purified F3 showed an absorbance spectrum characteristic of reaction centers, and contained 3.4 bacteriochlorophylls, 2.0 bacteriopheophytins, and 1.9 acid-labile iron atoms, but no cytochrome or ubiquinone (C-DOC reaction center). It had a weight of 1.2 X 10(5) daltons, and the main components were 4 protein species with molecular weights of 2.8 X 10(4), 2.7 X 10(4), 2.6 X 10(4), and 1.0 X 10(4). 6. The purified F4 showed a molecular weight of about 11,000, and contained one mole of ubiquinone-10 per mole (ubiquinone-10 protein). 7. The reaction center activity of C-DOC reaction centers was stimulated by ubiquinone-10 protein. In addition, the reaction center oxidized reduced cytochrome c2 in the light, provided that ubiquinone-10 protein was present (photo-oxidase activity).     

Murine alpha-fetoprotein: N-terminal amino acid sequence and C-terminal residue.

We have purified to homogeneity murine alpha-fetoprotein (MAFP) and determined the amino acid sequence of the first twenty-four residues. The N-terminal sequence obtained shows a high degree of homology with human and rat AFP's, but not human or rat albumins. The C-terminal residue is the same as human and “slow” rat AFP, but different from the corresponding albumins. We conclude that the AFP's are derived from homologous genes which are at best distantly related to the ancestral gene for albumin. The single C-terminal residue and N-terminal sequence suggests that the multiple forms of MAFP observed by others are due to carbohydrate micro-heterogeneity.