The influence of proteases on the activity of glucoamylase from Aspergillus niger C

Summary Two proteases from Aspergillus niger C post-culture medium were isolated by fractionation on a DEAE-sepharose column and ultrafiltration. The four fractions of glucoamylase activity (GA1, GA2, GA3 and GA4) present in the medium showed different susceptibility to the influence of proteases. The effects of proteases on the different glucoamylase fractions during the growth of the fungus are demonstrated. The activity was found to decrease at the beginning of the culture, but by its end there was a stimulation of GA4 glucoamylase. After treating GA2 and GA3 with protease II, a new additional fraction of glucoamylase was detected.     

Composition of Caulobacter crescentus murein synthesized in the presence of glycine

Abstract Glycine added to the growth medium of Caulobacter crescentus was found to substitute Cterminal alanine in the peptide side chains of the murein of this species. Murein synthesized in vivo and in vitro in the presence of glycerine was poorly crosslinked as was new murein formed in the presence of the amino acid. The reduced cross-linkage seems to be due to the effect of glycine on the formation of trimeric muropeptides as revealed by high-performance liquid chromatography (HPLC) muropeptide analysis of murein formed in the presence and absence of the amino acid.     

Biomimetic properties of simulated protomembranes

Origins of Life and Evolution of Biospheres -     

Primary structure differences of human surfactant-associated proteins isolated from normal and proteinosis lung.

The primary structures of human pulmonary surfactant-associated proteins SP-A, SP-B and SP-C isolated from lung lavage of patients with alveolar proteinosis exhibit significant differences from lung surfactant proteins isolated from lungs of healthy individuals. In contrast to SP-A from normal lungs, proteinosis SP-A was shown by SDS gel electrophoresis to contain large amounts of unreducibly cross-linked beta chains. Specific primary structure modifications of SP-C and SP-B proteins were established by direct molecular weight and structural analysis, using [252Cf]plasma desorption mass spectrometry (PD/MS) as the principal method. In comparison to normal lung surfactant SP-B, proteinosis SP-B showed a significantly increased molecular weight by approx. 500 Da for the unreduced protein dimer. SP-C proteins from normal lungs were identified to possess a bis-cysteinyl-5,6-(thioester)palmitoylated structure, and to contain a frayed N-terminus resulting in two sequences of 34 and 35 amino acid residues. In contrast, SP-C from proteinosis patients was modified by (i) partial or even complete removal of palmitate residues and (ii) additional N-terminal proteolytic degradation. These results indicate the presence of pathophysiological structure modifications, which are likely to occur in the alveolar space, and may lead to a reduced surfactant function.     

Structure and activity of the NAD(P)+‐dependent succinate semialdehyde dehydrogenase YneI from Salmonella typhimurium

Aldehyde dehydrogenases are found in all organisms and play an important role in the metabolic conversion and detoxification of endogenous and exogenous aldehydes. Genomes of many organisms including Escherichia coli and Salmonella typhimurium encode two succinate semialdehyde dehydrogenases with low sequence similarity and different cofactor preference (YneI and GabD). Here, we present the crystal structure and biochemical characterization of the NAD(P)⁺‐dependent succinate semialdehyde dehydrogenase YneI from S. typhimurium. This enzyme shows high activity and affinity toward succinate semialdehyde and exhibits substrate inhibition at concentrations of SSA higher than 0.1 mM. YneI can use both NAD⁺ and NADP⁺ as cofactors, although affinity to NAD⁺ is 10 times higher. High resolution crystal structures of YneI were solved in a free state (1.85 Å) and in complex with NAD⁺ (1.90 Å) revealing a two domain protein with the active site located in the interdomain interface. The NAD⁺ molecule is bound in the long channel with its nicotinamide ring positioned close to the side chain of the catalytic Cys268. Site‐directed mutagenesis demonstrated that this residue, as well as the conserved Trp136, Glu365, and Asp426 are important for activity of YneI, and that the conserved Lys160 contributes to the enzyme preference to NAD⁺. Our work has provided further insight into the molecular mechanisms of substrate selectivity and activity of succinate semialdehyde dehydrogenases. © 2012 Wiley Periodicals, Inc.     

Ecological causes of morphological evolution in the three‐spined stickleback

The central assumption of evolutionary theory is that natural selection drives the adaptation of populations to local environmental conditions, resulting in the evolution of adaptive phenotypes. The three‐spined stickleback (Gasterosteus aculeatus) displays remarkable phenotypic variation, offering an unusually tractable model for understanding the ecological mechanisms underpinning adaptive evolutionary change. Using populations on North Uist, Scotland we investigated the role of predation pressure and calcium limitation on the adaptive evolution of stickleback morphology and behavior. Dissolved calcium was a significant predictor of plate and spine morph, while predator abundance was not. Stickleback latency to emerge from a refuge varied with morph, with populations with highly reduced plates and spines and high predation risk less bold. Our findings support strong directional selection in three‐spined stickleback evolution, driven by multiple selective agents.     

The role of intraspecific competition in the dispersal of an invasive fish

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Potent V2 vasopressin antagonists with structural changes at their C-terminals

A variety of structural changes were made in the C-terminals of four potent antidiuretic (V2) antagonists. The parent analogs were all derivatives of [1-(beta-mercapto-beta,beta-cyclopentamethylenepropionic acid)]arginine-vasopressin, d(CH2)5AVP, namely d(CH2)5[D-Phe2,Ile4]AVP, d(CH2)5[D-Ile2,Ile4]AVP, d(CH2)5[D-Tyr(Et)2, Val4]AVP and d(CH2)5[D-Tyr(Et)2,Ile4]AVP. A number of amino acid amides were substituted for the C-terminal 9-glycinamide without reducing their V2-antagonistic potencies in rats. Many non-amino acid structures were also tolerated at the C-terminals of these antagonists and this end of these peptides can be prolonged without interfering with antagonistic potencies. Such altered V2-antagonists may be useful for the development of radioactive ligands, affinity labels and in affinity columns for studies on antidiuretic receptors. These C-terminal modifications also provide useful information for the further development of potent and specific V2-antagonists which can be valuable pharmacological tools and also promise to become useful clinically for the treatment of excessive water retention.     

The Feeding of Chaoborus flavicans Meigen (Diptera,Chaoboridae). and its Predation on Lake Zooplankton

In a study of Chaoborus feeding in a eutrophic lake, selectivity was found to be positive with Crustacea (especially copepodit stages). and negative with Rotatoria. Daily food rations were about 20% for most of the feeding period, but higher (106%). during the month of intensive growth after hatching. Feeding intensity correlated positively with amount of food an temperature, and negatively with Chaoborus concentration. Elimination of Crustacea (in the epilimnion of the central zone of the lake). equalled about 30–40% of Crustacea production in June and September and slightly exeeded the August production (it was almost zero in the remaining months because Chaoborus larvae stayed at the bottom). This applies, however, only in the central zone – about 50% of the lake volume. Chaoborus probably influences both the density of zooplankton and the quantitative relations between zooplankton species.     

Predicting transmembrane beta-barrels in proteomes

Very few methods address the problem of predicting beta-barrel membrane proteins directly from sequence. One reason is that only very few high-resolution structures for transmembrane beta-barrel (TMB) proteins have been determined thus far. Here we introduced the design, statistics and results of a novel profile-based hidden Markov model for the prediction and discrimination of TMBs. The method carefully attempts to avoid over-fitting the sparse experimental data. While our model training and scoring procedures were very similar to a recently published work, the architecture and structure-based labelling were significantly different. In particular, we introduced a new definition of beta- hairpin motifs, explicit state modelling of transmembrane strands, and a log-odds whole-protein discrimination score. The resulting method reached an overall four-state (up-, down-strand, periplasmic-, outer-loop) accuracy as high as 86%. Furthermore, accurately discriminated TMB from non-TMB proteins (45% coverage at 100% accuracy). This high precision enabled the application to 72 entirely sequenced Gram-negative bacteria. We found over 164 previously uncharacterized TMB proteins at high confidence. Database searches did not implicate any of these proteins with membranes. We challenge that the vast majority of our 164 predictions will eventually be verified experimentally. All proteome predictions and the PROFtmb prediction method are available at http://www.rostlab.org/ services/PROFtmb/.