Dark inhibition of ribulose-1,5-bisphosphate carboxylase/oxygenase in legumes: A biosystematic study

2-carboxyarabinitol 1-phosphate (CA 1-P) is a naturally occurring inhibitor of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco). Members of the Fabaceae exhibit a particularly wide range in the extent of CA 1-P accumulation during darkness and include Phaseolus vulgaris, whose dark/light regulation of Rubisco activity is principally achieved by synthesis/degradation of CA 1-P. An extensive survey of the degree of dark inhibition of Rubisco was undertaken for the subfamily Papilionoideae to elucidate evolutionary patterns in the occurrence of this regulatory mechanism. Seventy-five species from 21 tribes were examined. Dark inhibition of Rubisco was found in ancestral tribes such as the Sophoreae, but was substantially reduced or absent in representative species of three more recently evolved tribes, Cicereae, Hedysareae and Vicieae. We conclude that regulation of Rubisco by CA 1-P is neither of recent origin nor of restricted distribution among the Papilionoideae. On the contrary, it becomes lost or less pronounced only in a minority of the more evolutionarily advanced species in this important subfamily.     

Structural requirements for the binding of AMP to the allosteric site of NAD-specific isocitrate dehydrogenase from bakers' yeast

The specificity of yeast NAD-specific isocitrate dehydrogenase for the structures of the allosteric effector 5'-AMP was examined with analogues modified in the purine ring, pentosyl group, and 5'-phosphate group. An unsubstituted 6-amino group was essential for activation as was the phosphoryl group at the 5'-position. Activity was retained when an oxygen function of the 5'-phosphoryl was replaced by sulfur (Murry & Atkinson, 1968) or by nitrogen (phosphoramidates). 2-NH2-AMP, 2-azido-AMP, and 8-NH2-AMP were active; 8-azido-AMP and 8-Br-AMP were inactive. The configuration or nature of substituents about carbons 2' and 3' of the pentosyl portion of AMP was not critical for allosteric activation since AMP analogues containing, e.g., 2',3'-dideoxyribose or the bulky 2',3'-O-(2,4,6-trinitrocyclo-hexadienylidene) substituent (TNP-AMP) were active. TNP-AMP was bound to the enzyme with fluorescence enhancement and had an S0.5 for activation similar to the S0.5 for AMP. Positive effector activity was decreased when the pentosyl moiety of 5'-AMP was replaced by the six-membered nitrogen-containing morpholine group, indicating that the pentosyl group may be critical as a spacer for the proper geometry of binding to enzyme at the 6-amino and 5'-phosphoryl groups of 5'-AMP. A comparison of molecular models of 5'-AMP with 8,5'-cycloAMP suggests that the species of 5'-AMP required for binding to the enzyme contains the purine and ribose moieties in an anti conformation and positioning of the 5'-phosphate trans with respect to carbon 4'.(ABSTRACT TRUNCATED AT 250 WORDS)     

The photoreaction center of Rhodospirillum rubrum mutant strain F24.1

Rhodospirillum rubrum strain F24.1 is a spontaneous revertant of nonphototrophic mutant F24 derived from wild-type strain S1. Strain F24 shows no detectable photochemical activity and contains, at most, traces of the photoreaction center polypeptides. Strain F24.1 has a phototrophic growth rate close to that of the wild-type strain (Picorel, R., del Valle-Tascón, S. and Ramírez, J.M. (1977) Arch. Biophys. Biochem. 181, 665–670) but shows little photochemical activity. Light-induced absorbance changes in the near-infrared, photoinduced EPR signals and ferricyanide-elicited absorbance changes indicate that strain F24.1 has a photoreaction center content of 7–8% as compared to strain S1. Polyacrylamide gel electrophoresis of isolated F24.1 chromatophores shows the photoreaction center polypeptides to be present in amounts compatible with this value. Photoreaction center was prepared from strain F24.1 and showed no detectable difference with that of strain S1. It is concluded that strain F24.1 photosynthesis is due entirely to its residual 7–8% of typical photoreaction center.     

Role of angiotensin II and oxidative stress on renal aquaporins expression in hypernatremic rats

The aim of this study was to assess whether endogenous Ang II and oxidative stress produced by acute hypertonic sodium overload may regulate the expression of aquaporin-1 (AQP-1) and aquaporin-2 (AQP-2) in the kidney. Groups of anesthetized male Sprague–Dawley rats were infused with isotonic saline solution (control) or with hypertonic saline solution (Na group, 1 M NaCl), either alone or with losartan (10 mg kg^?1) or tempol (0.5 mg min^?1 kg^?1) during 2 h. Renal function parameters were measured. Groups of unanesthetized animals were injected intraperitoneally with hypertonic saline solution, with or without free access to water intake, Na+W, and Na?W, respectively. The expression of AQP-1, AQP-2, Ang II, eNOS, and NF-kB were evaluated in the kidney by Western blot and immunohistochemistry. AQP-2 distribution was assessed by immunofluorescence. Na group showed increased natriuresis and diuresis, and Ang II and NF-kB expression, but decreased eNOS expression. Losartan or tempol enhanced further the diuresis, and AQP-2 and eNOS expression, as well as decreased Ang II and NF-kB expression. Confocal immunofluorescence imaging revealed labeling of AQP-2 in the apical plasma membrane with less labeling in the intracellular vesicles than the apical membrane in kidney medullary collecting duct principal cells both in C and Na groups. Importantly, our data also show that losartan and tempol induces a predominantly accumulation of AQP-2 in intracellular vesicles. In unanesthetized rats, Na+W group presented increased diuresis, natriuresis, and AQP-2 expression (112?±?25 vs 64?±?16; *p?<?0.05). Water deprivation increased plasma sodium and diuresis but decreased AQP-2 (46?±?22 vs 112?±?25; §p?<?0.05) and eNOS expression in the kidney. This study is a novel demonstration that renal endogenous Ang II–oxidative stress, induced in vivo in hypernatremic rats by an acute sodium overload, regulates AQP-2 expression.     

Trypanosoma cruzi: Insights into naphthoquinone effects on growth and proteinase activity

In this study we compared the effects of naphthoquinones (α-lapachone, β-lapachone, nor-β-lapachone and Epoxy-α-lap) on growth of Trypanosoma cruzi epimastigotes forms, and on viability of VERO cells. In addition we also experimentally analyzed the most active compounds inhibitory profile against T. cruzi serine- and cysteine-proteinases activity and theoretically evaluated them against cruzain, the major T. cruzi cysteine proteinase by using a molecular docking approach. Our results confirmed β-lapachone and Epoxy-α-lap with a high trypanocidal activity in contrast to α-lapachone and nor-β-lapachone whereas Epoxy-α-lap presented the safest toxicity profile against VERO cells. Interestingly the evaluation of the active compounds effects against T. cruzi cysteine- and serine-proteinases activities revealed different targets for these molecules. β-Lapachone is able to inhibit the cysteine-proteinase activity of T. cruzi proteic whole extract and of cruzain, similar to E-64, a classical cysteine-proteinase inhibitor. Differently, Epoxy-α-lap inhibited the T. cruzi serine-proteinase activity, similar to PMSF, a classical serine-proteinase inhibitor. In agreement to these biological profiles in the enzymatic assays, our theoretical analysis showed that E-64 and β-lapachone interact with the cruzain specific S2 pocket and active site whereas Epoxy-α-lap showed no important interactions. Overall, our results infer that β-lapachone and Epoxy-α-lap compounds may inhibit T. cruzi epimastigotes growth by affecting T. cruzi different proteinases. Thus the present data shows the potential of these compounds as prototype of protease inhibitors on drug design studies for developing new antichagasic compounds.