Cathepsin B and D activity in stimulated peritoneal macrophages

Summary Highly sensitive and specific synthetic substrates were used to quantitate cathepsin B and D activity in peritoneal macrophages in response to stimulation in vivo with mineral oil and thioglycollate. After intraperitoneal instillation of mineral oil the activity of cathepsin B increased significantly (to 15 300 units/mg protein versus 7 340 in saline controls), reaching values approaching those found in alveolar macrophages (18 400 units/mg protein). Significantly greater stimulation of enzyme activity was obtained after intraperitoneal instillation of thioglycollate (23 600 units/mg protein). Cathepsin D activity also increased significantly after both mineral oil and thioglycollate. However, the increase was moderate (from 806 to about 1 200 units/mg protein), remaining still more than six times lower-than in alveolar macrophages. The data are the first to demonstrate that cathepsin B activity can be stimulated in vivo in peritoneal macrophages by instillation of agents that induce acute inflammation. They also point to a differential control of expression of cathepsin B and D activity in both peritoneal and alveolar macrophages in spite of the common lysosomal origin of the two enzymes.Abbreviations Cbz -N-benxyloxycarbonyl - 2NA 2-naphthylamine - EDTA ethylenediamine tetraacetate - DMSO dimethylsulfoxide - PBS phosphate-buffered saline - PM peritoneal macrophage - AM alveolar macrophage     

The impact of grazing on plant communities,plant populations and soil conditions on salt marshes

Grazing an abandoned salt marsh causes retrogressive succession, since mid salt-marsh communities change into lower salt-marsh communities. Grazing and mowing are compared in detail. Both management practices enhance species diversity in an abandoned salt marsh. This can be attributed to the removal of litter. The finding that lower salt-marsh species appear more with grazing than with mowing or abandoning is not related to a higher soil salinity as compared to mowing or abandoning, but probably to locally baring of the soil by grazing animals. Only species of pioneer or unstable environments seem to have a persistent seed bank, for other species seed dispersal seems to be a limiting factor for their establishment.Nomenclature follows Heukels & van Ooststroom (1977) for species; Westhoff & den Held (1969) for syntaxa.Mrs R. Rusthoven analyzed the soil samples, Mr E. Leeuwinga drawed the figures, and Mrs J. O'Brien corrected the English text.     

Genes Affecting the Regulation of SUC2 Gene Expression by Glucose Repression in SACCHAROMYCES CEREVISIAE

Mutants of Saccharomyces cerevisiae with defects in sucrose or raffinose fermentation were isolated. In addition to mutations in the SUC2 structural gene for invertase, we recovered 18 recessive mutations that affected the regulation of invertase synthesis by glucose repression. These mutations included five new snf1 (sucrose nonfermenting) alleles and also defined five new complementation groups, designated snf2, snf3, snf4, snf5, and snf6. The snf2, snf4, and snf5 mutants produced little or no secreted invertase under derepressing conditions and were pleiotropically defective in galactose and glycerol utilization, which are both regulated by glucose repression. The snf6 mutant produced low levels of secreted invertase under derepressing conditions, and no pleiotropy was detected. The snf3 mutants derepressed secreted invertase to 10-35% the wild-type level but grew less well on sucrose than expected from their invertase activity; in addition, snf3 mutants synthesized some invertase under glucose-repressing conditions.--We examined the interactions between the different snf mutations and ssn6, a mutation causing constitutive (glucose-insensitive) high-level invertase synthesis that was previously isolated as a suppressor of snf1. The ssn6 mutation completely suppressed the defects in derepression of invertase conferred by snf1, snf3, snf4 and snf6, and each double mutant showed the constitutivity for invertase typical of ssn6 single mutants. In contrast, snf2 ssn6 and snf5 ssn6 strains produced only moderate levels of invertase under derepressing conditions and very low levels under repressing conditions. These findings suggest roles for the SNF1 through SNF6 and SSN6 genes in the regulation of SUC2 gene expression by glucose repression.     

Sequence of Guinea Pig Myelin Basic Protein

This paper proposes a tentative amino acid sequence of guinea pig myelin basic protein obtained by comparison of peptide fragments of the guinea pig and bovine proteins. Analyses of the tryptic peptides confirmed the known sequence differences in the NH2-terminal half of the molecule and showed that in the COOH-terminal half of the guinea pig protein Ser131 was missing, Ala136 - His137 was deleted, Leu140 was replaced by Phe, and an extra Ala was inserted somewhere within sequence 142-151 (tryptic peptide T23 ). Sequence determination of guinea pig tryptic peptides corresponding to residues 130-134 ( T20 ), 135-138 ( T21 ), and 142-151 ( T23 ) of the bovine protein confirmed the above sequence changes and placed the extra Ala between Gly142 and His143 . The sequence of the region corresponding to bovine residues 130-143 is thus Ala-Asp-Tyr-Lys-Ser-Lys-Gly-Phe-Lys-Gly-Ala-His. No species differences were observed in the amino acid compositions of the remaining tryptic peptides obtained from the COOH-terminal half of the molecule. Based upon these results, the guinea pig basic protein contains 167 amino acid residues and has a molecular weight of 18,256.     

Correspondence of the monocyte antigen HMA-1 to the non-HLA antigen 9a

Monocyte-specific antibodies are detrimental to bone marrow and renal transplantation. By using human antimonocyte sera we were able to identify two monocyte-specific antigens, human monocyte antigen 1 and 2 (HMA-1 and HMA-2). The presence of HMA-1 and HMA-2 was compared with the presence of several non-HLA antigens. In panel and inhibition studies, HMA-1 corresponded to the previously described non-HLA granulocyte antigen 9a. Absorption studies showed that HMA-1 and 9a were both present on granulocytes and monocytes. The clinical relevance of these antigens is discussed.     

Regulation and structure of an Escherichia coli gene coding for an outer membrane protein involved in export of K88ab fimbrial subunits.

The nucleotide sequence of the faeD gene of Escherichia coli and the amino acid sequence of its product is presented. The faeD product is an outer membrane protein required for transport of K88ab fimbrial subunits across the outer membrane. The protein is synthesized as a precursor containing a signal peptide, and the tentative mature protein comprises 777 amino acid residues. The distribution of amino acids in the faeD protein is similar to that of other outer membrane proteins; showing a fairly even distribution of charged residues and the absence of extensive hydrophobic stretches. Secondary structure predictions revealed a region of 250 amino acid residues which might be embedded in the outer membrane. The 5'-end of faeD is located within a region showing dyad symmetry. This region serves to couple translation of faeD to the translation of the gene preceding it (faeC). The 3'-end of faeD shows an overlap of 5 bases with the next gene (faeE).     

Trapping, loss and annihilation of excitations in a photosynthetic system: II. Experiments with the purple bacteria Rhodospirillum rubrum and Rhodopseudomonas capsulata

In this paper a number of experiments with the purple bacteria Rhodospirillum rubrum and Rhodopseudomonas capsulata is described in which the total fluorescence yield and/or the total fraction of reaction centers closed after a picosecond laser pulse were measured as a function of the pulse intensity. The conditions were such that the reaction centers were either all in the open or all in the closed state before the pulse arrived. These experiments are analysed using the theoretical formalism discussed in the preceding paper (Den Hollander, W.T.F., Bakker J.G.C., and Van Grondelle, R., Biochim. Biophys. Acta 725, 492–507). From the experimental results the number of connected photosynthetic units, λ, the rate of energy transfer between neighboring antenna molecules, k_h, and the rate of trapping by an open reaction center, k^o_t, can be estimated. For R. rubrum it is found that λ = 14−17, k_h = (1−2)·10¹² s⁻¹ and k^o_t = (4−6)·10¹¹ s⁻¹, for Rps. capsulata λ ≈ 30, k_h ≈ 4·10¹¹ s⁻¹ and k^o_t ≈ 3·10¹¹ s⁻¹. The findings are discussed in terms of current models for the structure of the antenna and the kinetic properties of the decay processes occurring in these purple bacteria.     

ELECTROPHORETIC CHARACTERIZATION OF BASIC PROTEINS IN ACID EXTRACTS OF CENTRAL NERVOUS SYSTEM TISSUE

A technique has been outlined for identification of myelin basic proteins in mixtures of CNS proteins. Myelin basic proteins can be recognized easily by high cathodic mobility at low pH, a unique electrophoretic pattern exhibited at high pH and a characteristic colour when complexed with Amido black. The major protein extracted at pH 3·0 from either brain or spinal cord is myelin basic protein. In the low pH electrophoretic pattern of these extracts it is the most conspicuous component and the component migrating farthest cathodically; it does not appear in comparable electrophoretic patterns of liver extracts. Guinea pig myelin basic protein appears as a single dense blue-green band in low pH electrophoretic patterns, in contrast to the other proteins which are stained greyish-blue or greyish-purple by Amido black. The pattern of rat myelin basic protein is similar except that it consists of a pair of dense blue-green bands. A third characteristic which facilitates the identification of myelin basic proteins in mixtures is a considerable cathodic mobility and electrophoretic heterogeneity at pH 10·6. Most other basic CNS proteins barely penetrate the gel at this pH. We have also examined in detail the behaviour of two other components of pH 3·0 extracts which migrate close to myelin basic protein at low pH. Both are present in pH 3·0 extracts of liver and brain but not of spinal cord, and both stain grey instead of blue-green, a characteristic which readily distinguishes them from myelin basic protein. Neither of these components affects the characteristic pattern of microheterogeneity observed in high pH electrophoretograms of myelin basic proteins. One of these components has been purified and tentatively identified as lysine-rich histone F1.     

Repair of Alkylated Bacteriophage T4 Deoxyribonucleic Acid by a Mechanism Involving Polynucleotide Ligase

Methyl methanesulfate-induced lesions in bacteriophage T4 are repaired primarily by a mechanism involving polynucleotide ligase. Apparently, other recombinational and ultraviolet repair functions aren't involved.