Crystal structures of two homologous pathogenesis-related proteins from yellow lupine

Pathogenesis-related class 10 (PR10) proteins are restricted to the plant kingdom where they are coded by multigene families and occur at high levels. In spite of their abundance, their physiological role is obscure although members of a distantly related subclass (cytokinin-specific binding proteins) are known to bind plant hormones. PR10 proteins are of special significance in legume plants where their expression patterns are related to infection by the symbiotic, nitrogen-fixing bacteria. Here we present the first crystal structures of classic PR10 proteins representing two homologues from one subclass in yellow lupine. The general fold is similar and, as in a birch pollen allergen, consists of a seven-stranded beta-sheet wrapped around a long C-terminal helix. The mouth of a large pocket formed between the beta-sheet and the helix seems a likely site for ligand binding. The shape of the pocket varies because, in variance with the rigid beta-sheet, the helix shows unusual conformational variability consisting in bending, disorder, and axial shifting. A surface loop, proximal to the entrance to the internal cavity, shows an unusual structural conservation and rigidity in contrast to the high glycine content in its sequence. The loop is different from the so-called glycine-rich P-loops that bind phosphate groups of nucleotides, but it is very likely that it does play a role in ligand binding in PR10 proteins.     

Sequence of a novel plant ras-related cDNA from Pisum sativum

A clone isolated from a purple podded pea (Pisum sativum L.) cDNA library was shown to contain the complete coding sequence of a polypeptide with considerable homology to various members of the ras superfamily. The ras superfamily are a group of monomeric GTP-binding proteins of 21–25 kDa found in eukaryotic cells. Conserved sequences in the isolated clone include the GTP-binding site, GDP/GTP hydrolysis domain and C-terminal Cys residues involved in membrane attachment. Comparisons of the predicted amino acid sequence with those of other ras proteins show significantly higher homologies (ca. 70%) to two mammalian gene products, those of the BRL-ras oncogene, and the canine rab7 gene, than to any of the plant ras gene products so far identified (<40% homology). The high percentage of amino acid identity suggests that this cDNA may be the product of a gene, designated Psa-rab, which is the plant counterpart of rab7. Rab/ypt proteins are a subfamily of the ras superfamily thought to be involved in intracellular transport from the endoplasmic reticulum to the Golgi apparatus and in vesicular transport.Northern blot hybridisation analysis of total RNA from green and purple podded pea revealed a mRNA species of approximately the same size as the isolated cDNAs.     

The effects of dietary chromium supplementation on some blood parameters in sheep

The effects of dietary inorganic chromium on some biochemical parameters were determined in lambs fed either a control diet or a 200-ppb or 400-ppb chromium-supplemented diet. The live weight of the animals were measured and jugular blood samples were collected prior to supplementation (d 0) and on d 20, 40, and 55. On d 55, three animals from each group were slaughtered to measure subcutaneous fat. Sera were analyzed for glucose, triglyceride, total cholesterol, high-density lipoprotein (HDL) and low-density lipoprotein (LDL) cholesterol, total protein, albumin, ALT, AST, and GGT levels. Chromium supplementation had no significant effect on live weight, but subcutaneous fat was reduced significantly in both chromium groups. There was a slight decrease in glucose concentrations in the 200-ppb chromium group, although only the differences on d 55 were significant. Triglyceride levels in both chromium groups were lower than the control group with marked differences in the 400-ppb chromium group. HDL cholesterol levels increased in both treatment groups compare to control, although the differences in the 400-ppb chromium group on d 40 were significant. Serum Cr concentrations slightly but not significantly increased in both chromium groups. No significant differences were found in total and LDL cholesterol, total protein, albumin, ALT, AST, and GGT levels. In conclusion, chromium supplementation may affect carbohydrate and lipid metabolisms and lipid deposition in lambs.     

Recognition and degradation of chitin by streptomycetes

Chitin is the second most abundant renewable polysaccharide, as it is a component of the exoskeleton of many organisms and of the cell walls of numerous fungi. Most streptomycetes secrete a number of chitinases, hydrolyzing chitin to oligomers, chitobiose or N-acetylglucosamine which can be utilized as carbon or nitrogen source. The chitinases of several streptomycetes have been shown to have a modular arrangement comprising catalytic, substrate binding as well as linker domains. Moreover, during growth in the presence of chitin-containing substrates, many Streptomycesstrains have been shown to secrete formerly unknown, small (about 200 aa) chitin binding proteins (CHBs) which lack enzymatic activity and specifically target and invade chitin. Several motifs, including the relative location and spacing of four tryptophan residues, are conserved in the investigated CHB types, CHB1 and CHB2. The affinity of CHB1 to crab shell chitin is two times higher than that of CHB2. Comparative studies of various generated mutant CHB1 proteins led to the conclusion that it is one of the exposed tryptophan residues that directly contributes to the interaction with chitin. On the basis of immunological, biochemical and physiological studies, it can be concluded that the CHBs act like a glue with which streptomycetes target chitin-containing samples or organisms. The ecological implications of these findings are discussed.     

Peptidase N of Pseudomonas aeruginosa

Abstract Pseudomonas aeruginosa possesses a peptidase N activity analogous to those described in Escherichia coli and Salmonella typhimurium . This activity resides in a protein with an M _r value of 85 000. Part of this peptidase activity appears to be associated with the cytoplasmic membrane. The K _M value for this peptidase bound to the cytoplasmic membrane is in the range of 0.5 mM.     

Dihydropyridine Modulation of Voltage-Activated Calcium Channels in PC12 Cells: Effect of Pertussis Toxin Pretreatment

In this study, we report the effect of pertussis toxin pretreatment on dihydropyridine modulation of voltage-sensitive calcium channels in PC12 cells. The rise in intracellular calcium concentration caused by potassium depolarization is not affected significantly by pertussis toxin pretreatment. Nicardipine, a dihydropyridine derivative, added either before or after potassium-induced depolarization, reduces the resultant elevation in cytosolic calcium level both in control and in pertussis toxin-treated cells. The dihydropyridine agonist Bay K 8644, when added before potassium, is able to enhance the potassium-induced spike of cytosolic calcium levels, an effect significantly reduced by pertussis toxin pretreatment. Moreover, the addition of Bay K 8644 after potassium holds the intracellular calcium concentration at a cytosolic sustained level during the slow inactivating phase of depolarization. This effect of Bay K 8644 is inhibited by nicardipine. Pertussis toxin pretreatment slightly weakens the effect of Bay K 8644 when added after potassium-induced depolarization, whereas it significantly reduces the nicardipine inhibition of cytosolic calcium rise stimulated by potassium and Bay K 8644, but not by potassium alone. In conclusion, our findings suggest that a pertussis toxin-sensitive guanine nucleotide regulatory protein could be involved in the interaction between dihydropyridine derivatives and voltage-dependent calcium channels.     

Vacuolar sorting determinants within a plant storage protein trimer act cumulatively

The mechanism for vacuolar sorting of seed storage proteins is as yet poorly understood and no receptor has been identified to date. The homotrimeric glycoprotein phaseolin, which is the major storage protein of the common bean, requires a transient tetrapeptide at the C-terminus for its vacuolar sorting. A mutated construct without the tetrapeptide is secreted. We show here that coexpression of wild-type phaseolin and the mutated, secreted form in transgenic tobacco results in the formation of mixed trimers and partial vacuolar delivery of the mutated polypeptides and partial secretion of wild-type polypeptides. This indicates that the sorting signal has a cumulative effect within a phaseolin trimer. The result is discussed in the light of the hypothesized mechanisms for vacuolar sorting of seed storage proteins.     

Peptide-mediated DNA condensation for non-viral gene therapy

The construction of non-viral, virus-like vehicles for gene therapy involves the functionalization of multipartite constructs with nucleic acid-binding, cationic agents. Short basic peptides, alone or as fusion proteins, are appropriate DNA binding and condensing elements, whose incorporation into gene delivery vehicles results in the formation of protein–DNA complexes of appropriate size for cell internalization and intracellular trafficking. We review here the most used cationic peptides for artificial virus construction as well as the recently implemented strategies to control the architecture and biological activities of the resulting nanosized particles.