Entropy Drives an Attached Water Molecule from the C- to N-Terminus on Protonated Proline

Results from infrared photodissociation (IRPD) spectroscopy and kinetics of singly hydrated, protonated proline indicate that the water molecule hydrogen bonds preferentially to the formally neutral carboxylic acid at low temperatures and at higher temperatures to the protonated N-terminus, which bears the formal charge. Hydration isomer populations obtained from IRPD kinetic data as a function of temperature are used to generate a van`t Hoff plot that reveals that C-terminal binding is enthalpically favored by 4.2-6.4 kJ/mol, whereas N-terminal binding is entropically favored by 31-43 J/(mol K), consistent with a higher calculated barrier for water molecule rotation at the C-terminus.

Changes in leaf proline and fruit production induced by potassium stress in eggplant

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Proline content and protein patterns in Eucalyptus grandis shoots submitted to high and low temperature shocks

As respostas às mudanças de temperatura de plantas aclimatadas e não aclimatadas de E. grandis cultivadas in vitro foram avaliadas considerando alterações dos níveis de prolina e proteínas solúveis totais. Análises de proteínas solúveis através de SDS-PAGE e prolina foram realizadas após 12h a 12ºC (aclimatação ao frio) ou a 33ºC (aclimatação ao calor), e imediatamente depois dos choques térmicos a 41ºC e 0ºC. Análises também foram realizadas após um período de 24h depois dos choques térmicos (período de recuperação). O tratamento de temperatura a 0ºC não alterou o padrão de proteínas nas plantas aclimatadas e não aclimatadas, entretanto a temperatura baixa induziu altos níveis de prolina, que se mantiveram relativamente altos após o período de recuperação. Três novas proteínas (90,5, 75 e 39 kDa), provavelmente HSPs, foram observadas nas plantas aclimatadas e não aclimatadas submetidas às temperaturas altas. As plantas expostas a 41ºC foram capazes de recuperar-se dos choques após o período de recuperação, entretanto não houve recuperação completa das plantas expostas às baixas temperaturas. O efeito da aclimatação sobre a recuperação (homeostasis) pode variar dependendo do parâmetro avaliado, tipo e duração do choque térmico.

Free proline accumulation in sugarcane under water restriction and spittlebug infestation

A cigarrinha-das-raízes Mahanarva fimbriolata (Stål) (Hemiptera: Cercopidae) tornou-se uma praga importante da cultura da cana-de-açúcar na região centro-sul do Brasil. Estratégias de controle têm sido desenvolvidas, mas os danos promovidos por essa praga e sua interação com a cana-de-açúcar ainda são pouco caracterizados. em altos níveis de infestação da praga, os sintomas nas plantas são muito semelhantes à restrição hídrica severa. Este trabalho foi conduzido com o objetivo de determinar se o estresse promovido pela infestação de ninfas de cigarrinha-das-raízes resulta em acúmulo de prolina livre, além de comparar dois genótipos de cana-de-açúcar quanto à tolerância ao déficit hídrico. Dois experimentos foram conduzidos em delineamento inteiramente casualizado, em esquema fatorial 2 x 2 x 4 com duas cultivares (SP80-1816 e RB72454), dois níveis de estresse (controle e dez ninfas por planta ou 50% de restrição hídrica) e quatro épocas. O déficit hídrico promovido por M. fimbriolata não resultou em acúmulo de prolina, ilustrando a existência de mecanismos distintos para a percepção do estresse hídrico promovido pela praga e pela variação no potencial osmótico nas células das raízes. A cultivar RB72454 acumula maiores teores de prolina livre, e o acúmulo de massa seca e o crescimento dos colmos são menos afetados nessa cultivar sob restrição hídrica. Os teores de solutos compatíveis provavelmente não podem ser utilizados para quantificar o estresse promovido por M. fimbriolata em cana-de-açúcar. A cultivar RB72454 é mais tolerante ao déficit hídrico do que a cultivar SP80-1816.

Catechol biosensing using a nanostructured layer-by-layer film containing Cl-catechol 1,2-dioxygenase

dThe detection of aromatic compounds from pesticides and industrial wastewater has become of great interest, since these compounds withstand chemical oxidation and biological degradation, accumulating in the environment. In this work, a highly sensitive biosensor for detecting catechol was obtained with the immobilization of Cl-catechol 1,2-dioxygenase (CCD) in nanostructured films. CCD layers were alternated with poly(amidoamine) generation 4 (PAMAM G4) dendrimer using the electrostatic layer-by-layer (LbL) technique. Circular dichroism (CD) measurements indicated that the immobilized CCD preserved the same conformation as in solution. The thickness of the very first CCD layers in the LbL films was estimated at ca. 3.6 nm, as revealed by surface plasmon resonance (SPR). PAMAM/CCD 10-bilayer films were employed in detecting diluted catechol solutions using either an optical or electrical approach. Due to the mild immobilization conditions employed, especially regarding the pH and ionic strength of the dipping solutions, CCD remained active in the films for periods longer than 3 weeks. The optical detection comprised absorption experiments in which the formation of cis-cis muconic acid, resulting from the reaction between CCD and catechol, was monitored by measuring the absorbance at 260 nm after film immersion in catechol solutions. The electrical detection was carried out using LbL films deposited onto gold-interdigitated electrodes immersed in aqueous solutions at different catechol concentrations. Using impedance spectroscopy in a broad frequency range (1Hz-1kHz), we could detect catechol in solutions at concentrations as low as 10(-10) M. (c) 2005 Elsevier B.V. All rights reserved.

On-line redox speciation analysis of antimony using L-proline immobilized on controlled pore glass and hydride generation inductively coupled plasma optical emission spectrometry for detection

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

Eucalyptus ESTs involved in the production of 9-cis epoxycarotenoid dioxygenase, a regulatory enzyme of abscisic acid production

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

Photosynthetic responses and proline content of mature and young leaves of sunflower plants under water deficit

In mature and young leaves of sunflower (Helianthus annuus L. cv. Catissol-01) plants grown in the greenhouse, photosynthetic rate, stomatal conductance, and transpiration rate declined during water stress independently of leaf age and recovered after 24-h rehydration. The intercellular CO 2 concentration, chlorophyll (Chl) content, and photochemical activity were not affected by water stress. However, non-photochemical quenching increased in mature stressed leaves. Rehydration recovered the levels of non-photochemical quenching and increased the F v/F m in young leaves. Drought did not alter the total Chl content. However, the accumulation of proline under drought was dependent on leaf age: higher content of proline was found in young leaves. After 24 h of rehydration the content of proline returned to the same contents as in control plants.

Indoleamine 2,3-dioxygenase (IDO) Activity in Placental Compartments of Renal-Transplanted Pregnant Women

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

P-I class metalloproteinase from Bothrops moojeni venom is a post-proline cleaving peptidase with kininogenase activity: Insights into substrate selectivity and kinetic behavior

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Cardiovascular effects of the microinjection of L-proline into the third ventricle or the paraventricular nucleus of the hypothalamus in unanesthetized rats

We investigated the cardiovascular effects of the microinjection of L-proline (L-Pro) into the third ventricle (3V) and its peripheral mechanisms. Different doses of L-Pro into the 3V caused dose-related pressor and bradycardiac responses. The pressor response to L-Pro injected into the 3V was potentiated by intravenous pretreatment with the ganglion blocker pentolinium (5 mg/kg), thus excluding any significant involvement of the sympathetic nervous system. Because the response to the microinjection of L-Pro into the 3V was blocked by intravenous pretreatment with the V1-vasopressin receptor antagonist dTyr(CH2)5(Me)AVP (50 mu g/kg), it is suggested that these cardiovascular responses are mediated by a vasopressin release. The pressor response to the microinjection of L-Pro into the 3V was found to be mediated by circulating vasopressin, so, given that the paraventricular nucleus of the hypothalamus (PVN) is readily accessible from the 3V, we investigated whether the PVN could be a site of action for the L-Pro microinjected in the 3V. The microinjection of L-Pro (0.033 mu moles/0.1 mu l) into the PVN caused cardiovascular responses similar to those of injection of the 3V and were also shown to be mediated by vasopressin release. In conclusion, these results show that the microinjection of L-Pro into the 3V causes pressor and bradycardiac responses that could involve stimulation of the magnocellular cells of the PVN and release of vasopressin into the systemic circulation. Also, because the microinjection of L-Pro into the PVN caused a pressor response, this is the first evidence of cardiovascular effects caused by its injection in a supramedullary structure. (c) 2012 Wiley Periodicals, Inc.

Exogenous ornithine is an effective precursor and the delta-ornithine amino transferase pathway contributes to proline accumulation under high N recycling in salt-stressed cashew leaves

The role of the delta-ornithine amino transferase (OAT) pathway in proline synthesis is still controversial and was assessed in leaves of cashew plants subjected to salinity. The activities of enzymes and the concentrations of metabolites involved in proline synthesis were examined in parallel with the capacity of exogenous ornithine and glutamate to induce proline accumulation. Proline accumulation was best correlated with OAT activity, which increased 4-fold and was paralleled by NADH oxidation coupled to the activities of OAT and Delta(1)-pyrroline-5-carboxylate reductase (P5CR), demonstrating the potential of proline synthesis via OAT/P5C. Overall, the activities of GS. GOGAT and aminating GDH remained practically unchanged under salinity. The activity of P5CR did not respond to NaCl whereas Delta(1)-pyrroline-5-carboxylate dehydrogenase was sharply repressed by salinity. We suggest that if the export of P5C from the mitochondria to the cytosol is possible, its subsequent conversion to proline by P5CR may be important. In a time-course experiment, proline accumulation was associated with disturbances in amino acid metabolism as indicated by large increases in the concentrations of ammonia, free amino acids, glutamine, arginine and ornithine. Conversely, glutamate concentrations increased moderately and only within the first 24 h. Exogenous feeding of ornithine as a precursor was very effective in inducing proline accumulation in intact plants and leaf discs, in which proline concentrations were several times higher than glutamate-fed or salt-treated plants. Our data suggest that proline accumulation might be a consequence of salt-induced increase in N recycling, resulting in increased levels of ornithine and other metabolites involved with proline synthesis and OAT activity. Under these metabolic circumstances the OAT pathway might contribute significantly to proline accumulation in salt-stressed cashew leaves. (C) 2011 Elsevier GmbH. All rights reserved.

Effects of overinflation on procollagen type III expression in experimental acute lung injury

Abstract Introduction In acute lung injury (ALI), elevation of procollagen type III (PC III) occurs early and has an adverse impact on outcome. We examined whether different high-inflation strategies of mechanical ventilation (MV) in oleic acid (OA) ALI alter regional expression of PC III. Methods We designed an experimental, randomized, and controlled protocol in which rats were allocated to two control groups (no injury, recruited [alveolar recruitment maneuver after tracheotomy without MV; n = 4 rats] and control [n = 5 rats]) or four injured groups (one exposed to OA only [n = 10 rats] and three OA-injured and ventilated). The three OA-injured groups were ventilated for 1 hour according to the following strategies: LVHP-S (low volume-high positive end-expiratory pressure [PEEP], supine; n = 10 rats, tidal volume [VT] = 8 ml/kg, PEEP = 12 cm H2O), HVLP-S (high volume-low PEEP, supine; n = 10 rats, VT = 20 ml/kg, PEEP = 5 cm H2O), and HVLP-P (high volume-low PEEP, prone; n = 10 rats). Northern blot analysis for PC III and interleukin-1-beta (IL-1β) and polymorphonuclear infiltration index (PMI) counting were performed in nondependent and dependent regions. Regional differences between groups were assessed by two-way analysis of variance after logarithmic transformation and post hoc tests. Results A significant interaction for group and region effects was observed for PC III (p = 0.012) with higher expression in the nondependent region for HVLP-S and LVHP-S, intermediate for OA and HVLP-P, and lower for control (group effect, p < 0.00001, partial η2 = 0.767; region effect, p = 0.0007, partial η2 = 0.091). We found high expression of IL-1β (group effect, p < 0.00001, partial η2 = 0.944) in the OA, HVLP-S, and HVLP-P groups without regional differences (p = 0.16). PMI behaved similarly (group effect, p < 0.00001, partial η2 = 0.832). Conclusion PC III expression is higher in nondependent regions and in ventilatory strategies that caused overdistension. This response was partially attenuated by prone positioning.

Proline dehydrogenase regulates redox state and respiratory metabolism in Trypanosoma Cruzi

Over the past three decades, L-proline has become recognized as an important metabolite for trypanosomatids. It is involved in a number of key processes, including energy metabolism, resistance to oxidative and nutritional stress and osmoregulation. In addition, this amino acid supports critical parasite life cycle processes by acting as an energy source, thus enabling host-cell invasion by the parasite and subsequent parasite differentiation. In this paper, we demonstrate that L-proline is oxidized to Δ(1)-pyrroline-5-carboxylate (P5C) by the enzyme proline dehydrogenase (TcPRODH, E.C. 1.5.99.8) localized in Trypanosoma cruzi mitochondria. When expressed in its active form in Escherichia coli, TcPRODH exhibits a Km of 16.58±1.69 µM and a Vmax of 66±2 nmol/min mg. Furthermore, we demonstrate that TcPRODH is a FAD-dependent dimeric state protein. TcPRODH mRNA and protein expression are strongly upregulated in the intracellular epimastigote, a stage which requires an external supply of proline. In addition, when Saccharomyces cerevisiae null mutants for this gene (PUT1) were complemented with the TcPRODH gene, diminished free intracellular proline levels and an enhanced sensitivity to oxidative stress in comparison to the null mutant were observed, supporting the hypothesis that free proline accumulation constitutes a defense against oxidative imbalance. Finally, we show that proline oxidation increases cytochrome c oxidase activity in mitochondrial vesicles. Overall, these results demonstrate that TcPRODH is involved in proline-dependant cytoprotection during periods of oxidative imbalance and also shed light on the participation of proline in energy metabolism, which drives critical processes of the T. cruzi life cycle.