pH-Sensitive Binding of Cytochrome c to the Inner Mitochondrial Membrane. Implications for the Participation of the Protein in Cell Respiration and Apoptosis

Cytochrome c exhibits two positively charged sites: site A containing lysine residues with high pK(a) values and site L containing ionizable groups with pK(aobs),values around 7.0. This protein feature implies that cytochrome c can participate in the fusion of mitochondria and have its detachment from the inner membrane regulated by cell acidosis and alkalosis. In this study, We demonstrated that both horse and tuna cytochrome c exhibited two types of binding to inner mitochondrial membranes that contributed to respiration: a high-affinity and low-efficiency pi-I-independent binding (microscopic dissociation constant K(sapp2), similar to 10 nM) and a low-affinity and high-efficiency pH-dependent binding that for horse cytochrome c had a pK(a) of similar to 6.7. For tuna cytochrome c (Lys22 and His33 replaced with Asn and Trp, respectively), the effect of pH on K(sapp1), was less striking than for the horse heme protein, and both tuna and horse cytochrome c had closed K(sapp1) values at pH 7.2 and 6.2, respectively. Recombinant mutated cytochrome c H26N and H33N also restored the respiration of the cytochrome c-depleted mitoplast in a pH-dependent manner. Consistently, the detachment of cytochrome c from nondepleted mitoplasts was favored by alkalinization, suggesting that site Lionization influences the participation of cytochrome c in the respiratory chain and apoptosis.

The Xanthomonas citri effector protein PthA interacts with citrus proteins involved in nuclear transport, protein folding and ubiquitination associated with DNA repair

P>Xanthomonas axonopodis pv. citri utilizes the type III effector protein PthA to modulate host transcription to promote citrus canker. PthA proteins belong to the AvrBs3/PthA family and carry a domain comprising tandem repeats of 34 amino acids that mediates protein-protein and protein-DNA interactions. We show here that variants of PthAs from a single bacterial strain localize to the nucleus of plant cells and form homo- and heterodimers through the association of their repeat regions. We hypothesize that the PthA variants might also interact with distinct host targets. Here, in addition to the interaction with alpha-importin, known to mediate the nuclear import of AvrBs3, we describe new interactions of PthAs with citrus proteins involved in protein folding and K63-linked ubiquitination. PthAs 2 and 3 preferentially interact with a citrus cyclophilin (Cyp) and with TDX, a tetratricopeptide domain-containing thioredoxin. In addition, PthAs 2 and 3, but not 1 and 4, interact with the ubiquitin-conjugating enzyme complex formed by Ubc13 and ubiquitin-conjugating enzyme variant (Uev), required for K63-linked ubiquitination and DNA repair. We show that Cyp, TDX and Uev interact with each other, and that Cyp and Uev localize to the nucleus of plant cells. Furthermore, the citrus Ubc13 and Uev proteins complement the DNA repair phenotype of the yeast Delta ubc13 and Delta mms2/uev1a mutants, strongly indicating that they are also involved in K63-linked ubiquitination and DNA repair. Notably, PthA 2 affects the growth of yeast cells in the presence of a DNA damage agent, suggesting that it inhibits K63-linked ubiquitination required for DNA repair.

Structure, Dynamics, and RNA Interaction Analysis of the Human SBDS Protein

Shwachman-Bodian-Diamond syndrome is an autosomal recessive genetic syndrome with pleiotropic phenotypes, including pancreatic deficiencies, bone marrow dysfunctions with increased risk of myelodysplasia or leukemia, and skeletal abnormalities. This syndrome has been associated with mutations in the SBDS gene, which encodes a conserved protein showing orthologs in Archaea and eukaryotes. The Shwachman-Bodian-Diamond syndrome pleiotropic phenotypes may be an indication of different cell type requirements for a fully functional SBDS protein. RNA-binding activity has been predicted for archaeal and yeast SBDS orthologs, with the latter also being implicated in ribosome biogenesis. However, full-length SBDS orthologs function in a species-specific manner, indicating that the knowledge obtained from model systems may be of limited use in understanding major unresolved issues regarding SBDS function, namely, the effect of mutations in human SBDS on its biochemical function and the specificity of RNA interaction. We determined the solution structure and backbone dynamics of the human SBDS protein and describe its RNA binding site using NMR spectroscopy. Similarly to the crystal structures of Archaea, the overall structure of human SBDS comprises three well-folded domains. However, significant conformational exchange was observed in NMR dynamics experiments for the flexible linker between the N-terminal domain and the central domain, and these experiments also reflect the relative motions of the domains. RNA titrations monitored by heteronuclear correlation experiments and chemical shift mapping analysis identified a classic RNA binding site at the N-terminal FYSH (fungal, Yhr087wp, Shwachman) domain that concentrates most of the mutations described for the human SBDS. (C) 2010 Elsevier Ltd. All rights reserved.

Effects of Extrusion on the Emulsifying Properties of Rumen and Soy Protein

Defatted rumen protein and soy protein concentrate were extruded in a 15.5:1 L/D single-screw extruder at the optimum conditions for their expansion (150A degrees C and 35% moisture, and 130A degrees C and 35% moisture, respectively). Emulsions were produced with these proteins and studied by rheology and time domain low-resolution (1)H nuclear magnetic resonance (TD-NMR). Extrusion increased storage modulus of rumen protein emulsions. The opposite was observed for soy protein. Mechanical relaxation showed the existence of three relaxing components in the emulsions whose relative contributions were changed by extrusion. Likewise, spin-spin relaxation time constants (T (2)) measured by TD-NMR also showed three major distinct populations of protons in respect to their mobility that were also altered by extrusion. Extrusion increased surface hydrophobicity of both rumen and soy protein. Solubility of rumen protein increased with extrusion whereas soy protein had its solubility decreased after processing. Extrusion promoted molecular reorganization of protein, increasing its superficial hydrophobicity, affecting its interfacial properties and improving its emulsifying behavior. The results show that extrusion can promote the use of rumen, a by-product waste from the meat industry, in human nutrition by replacing soy protein in food emulsions.

Determination of crude protein, crude fiber and crude fat in twenty-five genotypes of kale (B-oleracea var. acephala)

The aim of the study was to determine the percentage of crude protein, crude fiber and crude fat (ether extract) of 25 genotypes of kale from the Germplasm Bank of Instituto Agronomico de Campinas and of one genotype grown in the region of Jaboticabal-SP. The plants were cultivated in the field, and the leaves after collection were pre-dried in a convection oven at 65 degrees C for 72 h. Afterward, the leaves were analyzed for crude protein, crude fiber and crude fat (ether-soluble materials). Significant differences were detected among the different genotypes for all the characteristics examined. of the genotypes studied, six showed more than 30% crude protein: HS-20 (32.56%), Comum (31.70%), Couve de Arthur Nogueira 2 (31.16%), Pires 2 de Campinas (30.63%), Manteiga 1-916 (30.36%), and Manteiga de Ribeirao Pires I-2446 (30.03%). In relation to crude fiber, the highest percentage was seen in the genotype Manteiga de Mococa (10.92%), differing significantly from the other genotypes studied. With regard to crude fat, the highest percentage was found in the genotype HS-20 (3.72%), and Pires 1 de Campinas (3.34%). of the genotypes tested, HS-20 stood out among the others, showing both the highest percentage of protein and fat.

Genetic parameters for milk, fat and protein yields in Murrah buffaloes (Bubalus bubalis Artiodactyla, Bovidae)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Prediction equations to estimate the demand of energy and crude protein for maintenance, gain and egg production for laying Japanese quails

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Molecular Modeling Suggests Cruzain Specificity for Peptide Primaquine Prodrugs

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Seasonal influence on biochemical profile and serum protein electrophoresis for Boa constrictor amarali in captivity

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

In vitro antibacterial activity of medicinal plant extracts against Escherichia coli strains from human clinical specimens and interactions with antimicrobial drugs

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Expression of renin-angiotensin system signalling compounds in maternal protein-restricted rats: effect on renal sodium excretion and blood pressure

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

In utero protein restriction causes growth delay and alters sperm parameters in adult male rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Recovery of rat growth and lipid profiles in adult rats subjected to fetal protein malnutrition with a fructose-rich diet

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Expression and purification of human respiratory syncytial virus recombinant fusion protein

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Protein, methionine+cystine and lysine levels for Japanese quails during the production phase

An experiment was conducted at Faculdade de Medicina Veterinária e Zootecnia/Unesp - Botucatu for 168 days to evaluate the effects of protein, Met + Cys and lysine diet levels on egg production and egg quality of laying Japanese quails. Quails with 42 days of age were reared in a completely randomized design. There were 1,944 quails distributed in four replicates of 27 birds per pen, according to a factorial 3x3x2 with three crude protein levels (16, 18 and 20% CP), three Met + Cys levels (0.700; 0.875 and 1.050%) and two lysine levels (1.100 and 1.375%). Birds fed diets with 18 and 20% CP had higher feed intake and egg production than those fed diets with 16% CP. There was significant interaction (p<0.05) between protein and Met + Cys levels on egg weight. There was no effects (p>0.05) of the protein level on feed conversion per dozen eggs; however, improved feed conversion per egg mass was seen for birds fed diets with 20% CP compared to those fed diets with 16% and 18% CP. Protein and lipid percentage in the yolk increased when dietary protein level increased from 16 to 18%. Increasing Met + Cys from 0.700% to 0.875% reduced yolk protein percentage. Higher lipid percentage in the yolk was seen in eggs from quails fed diets with 1.050% Met + Cys, whereas 1.375% lysine in the diet of resulted in decreased egg production and egg mass, besides poorer feed conversion per dozen eggs and per egg mass.