Towards Characterization of the Chloroplast NAD（P）H Dehydrogenase Complex

The NAD（P）H dehydrogenase （NDH） complex in chloroplast thylakoid membranes functions in cyclic electron transfer, and in chlororespiration. NDH is composed of at least 15 subunits, including both chloroplast- and nuclear-encoded proteins. During the past few years, extensive proteomic and genetic research on the higher plant NDH complex has been carried out, resulting in identification of several novel nuclear-encoded subunits. In addition, a number of auxiliary proteins, which mainly regulate the expression of chloroplast-encoded ndh genes as well as the assembly and stabilization of the NDH complex, have been discovered and characterized. In the absence of detailed crystallographic data, the structure of the NDH complex has remained obscure, and therefore the role of several NDH-associated nuclear-encoded proteins either as auxiliary proteins or structural subunits remains uncertain. In this review, we summarize the current knowledge on the subunit composition and assembly process of the chloroplast NDH complex. In addition, a novel oligomeric structure of NDH, the PSI/NDH supercomplex, is discussed.     

Sequence variation in CYP51A from the Y strain of Trypanosoma cruzi alters its sensitivity to inhibition

CYP51 (sterol 14α-demethylase) is an efficient target for clinical and agricultural antifungals and an emerging target for treatment of Chagas disease, the infection that is caused by multiple strains of a protozoan pathogen Trypanosoma cruzi. Here, we analyze CYP51A from the Y strain T. cruzi. In this protein, proline 355, a residue highly conserved across the CYP51 family, is replaced with serine. The purified enzyme retains its catalytic activity, yet has been found less susceptible to inhibition. These biochemical data are consistent with cellular experiments, both in insect and human stages of the pathogen. Comparative structural analysis of CYP51 complexes with VNI and two derivatives suggests that broad-spectrum CYP51 inhibitors are likely to be preferable as antichagasic drug candidates.     

Rainfall, phycocyanin, and N:P ratios related to cyanobacterial blooms in a Korean large reservoir

Nutrient concentrations and other environmental factors were measured in the Daechung Reservoir for 25 weeks from spring until autumn in 1999. The high irradiance after heavy rainfall provided optimal meteorological conditions for bloom formation during summer, therefore, rain would also appear to forecast imminent bloom. The bloom formation was largely governed by cyanobacteria, in particular, Microcystis spp. and Anabaenaspp. Phycocyanin showed higher correlation with cyanobacteria (r = 0.744, P < 0.001) compared to chlorophyll-a(r = 0.599, P < 0.01). Therefore, phycocyanin was more accurate and useful than chlorophyll-a in quantitatively measuring cyanobacterial blooms. The atomic N:P ratio of the particulate form also showed a high correlation with cyanobacteria (r = 0.541, P < 0.01), increasing from 4.3 to 14.6 during bloom formation, while that of the dissolved form decreased from 25.5 to 8.7. These results indicated that the algae assimilated N significantly without comparable P uptake during the blooming season, which was in sharp contrast to the excessive storage of P during the spring.     

Nutrient couplings between on-site sewage disposal systems, groundwaters, and nearshore surface waters of the Florida Keys

We performed a one-year study to determine the effects of on-site sewage disposal systems (OSDS, septic tanks) on the nutrient relations of limestone groundwaters and nearshore surface waters of the Florida Keys. Monitor wells were installed on canal residences with OSDS and a control site in the Key Deer National Wildlife Refuge on Big Pine Key. Groundwater and surface water samples were collected monthly during 1987 and analyzed for concentrations of dissolved inorganic nitrogen (DIN = NO_f3/sup- + NO_f2/sup- + NH_4/su+), soluble reactive phosphate (SRP), temperature and salinity. Significant nutrient enrichment (up to 5000-fold) occurred in groundwaters contiguous to OSDS; DIN was enriched an average of 400-fold and SRP some 70-fold compared to control groundwaters. Ammonium was the dominant nitrogenous species and its concentration ranged from a low of 0.77 μM in control wells to 2.75 mM in OSDS-enriched groundwaters. Concentrations of nitrate plus nitrite were also highly enriched and ranged from 0.05 μM in control wells to 2.89 mM in enriched groundwaters. Relative to DIN, concentrations of SRP were low and ranged from 30 nM in control wells to 107 μM in enriched groundwaters. N : P ratios of enriched groundwaters were consistently > 100 and increased with increasing distance from the OSDS, suggesting significant, but incomplete, adsorption of SRP by subsurface flow through carbonate substrata. Nutrient concentrations of groundwaters also varied seasonally and were approximately two-fold higher during the winter (DIN = 1035 μM; SRP = 10.3 μM) compared to summer (DIN = 470 μM; SRP = 4.0 μM). In contrast, surface water nutrient concentrations were two-fold higher during the summer (DIN = 5.0 μM; SRP = 0.50 μM) compared to winter (DIN = 2.5 μM; SRP = 0.15 μM). Direct measurement of subsurface groundwater flow rate indicated that tides and increased groundwater recharge enhanced flow some two-fold and six-fold, respectively. Accordingly, the observed seasonal coupling of OSDS-derived nutrients from groundwaters to surface waters is maximum during summer because of seasonally maximum tides and increased hydraulic head during the summer wet season. The yearly average benthic flux of anthropogenic DIN into contiguous canal surface waters is 55 mmol m^-2 day^-1, a value some five-fold greater than the highest rate of benthic N-fixation measured in carbonate-rich tropical marine waters.     

华山新麦草叶片的光合生理特性

对华山新麦草(Psathyrostachys huashanica Keng ex P.C.Kuo)营养叶的净光合速率(Pn)和蒸腾速率(Tr)的日变化曲线进行了分析,并在对叶温(Tl)、气孔阻力(Rs)、光合有效辐射强度(PAR)和气温(Ta)的日变化曲线进行测定的基础上分析了它们对华山新麦草Pn的影响规律。结果表明:华山新麦草Pn的日变化曲线呈"三峰"型,峰值分别为6.5、6.2和9.0μmol.m-2.s-1,依次出现在9:30、11:30和16:30,而且具有明显的"午降"现象;Tr的日变化曲线呈"单峰"型,最大值为1.7 mmol.m-2.s-1,出现在13:30;Tl、Rs、PAR和Ta的日变化曲线均呈"单峰"型,峰值分别出现在12:30、11:30、12:30和13:30。华山新麦草的Pn对Tl、PAR和Ta的响应曲线均呈"抛物线"型,Pn在一定范围内与Tl、PAR和Ta呈正相关,随着Tl、PAR和Ta的升高逐渐增加至最大值后逐渐降低;而Pn与Rs则呈负相关,Pn在一定范围内随Rs的增大逐渐降低。根据拟合方程,华山新麦草营养叶的光补偿点和光饱和点分别为1.1和531.5μmol.m-2.s-1,说明该种类具有很强的喜光性,且对光照强度的适应范围较广。研究结果表明:较大的气孔阻力是造成华山新麦草叶片净光合速率偏低的主要原因。     

Synchronous intra-Golgi transport induces the release of Ca from the Golgi apparatus

The mechanisms of secretory transport through the Golgi apparatus remain an issue of debate. The precise functional importance of calcium ions (Ca²⁺) for intra-Golgi transport has also been poorly studied. Here, using different approaches to measure free Ca²⁺ concentrations in the cell cytosol ([Ca²⁺]_cyt) and inside the lumen of the Golgi apparatus ([Ca²⁺]_GA), we have revealed transient increases in [Ca²⁺]_cyt during the late phase of intra-Golgi transport that are concomitant with a decline in the maximal [Ca²⁺]_GA restoration ability. Thus, this redistribution of Ca²⁺ from the Golgi apparatus into the cytosol during the movement of cargo through the Golgi apparatus appears to have a role in intra-Golgi transport, and mainly in the late Ca²⁺-dependent phase of SNARE-regulated fusion of Golgi compartments.     

Effects of Hypoglycaemia and Hypoxia on the Intracellular pH of Cerebral Tissue as Measured by 31P Nuclear Magnetic Resonance

Changes in high-energy phosphate metabolites and the intracellular pH (pHi) were monitored in cerebral tissue during periods of hypoglycaemia and hypoxia using 31P nuclear magnetic resonance spectroscopy. Superfused brain slices were loaded with deoxyglucose at a concentration shown not to impair cerebral metabolism, and the chemical shift of the resulting 2-deoxyglucose-6-phosphate (DOG6P) peak was used to monitor the pHi. In some experiments with low circulating levels of Pi, the intracellular Pi was visible and indicated a pH identical to that of DOG6P, an observation validating its use as an indicator of pHi in cerebral tissue. The pHi was found to be unchanged during moderate hypoglycaemia; however, mild hypoxia (PO2 = 16.4 kPa) and severe hypoglycaemia produced marked reductions from the normal of 7.2 to 6.8 and 7.0, respectively. Hypoglycaemia caused a fall in the level of both phosphocreatine (PCr) and ATP, whereas hypoxia affected PCr alone, as shown previously. However, the fall in pHi was similar during the two insults, thus indicating that the change in pH is not directly linked to lactate production or to the creatine kinase reaction.     

Evolution of the lipoprotein gene in the enterobacteriaceae. Cloning and DNA sequence of the lpp gene from Proteus mirabilis

We cloned the lipoprotein gene from Proteus mirabilis and determined its DNA sequence. Comparison with the lpp genes from Escherichia coli, Serratia marcescens, Erwinia amylovora and Morganella morganii revealed several unique features of the evolution of the lpp gene in the Enterobacteriaceae and enabled us to establish phylogenetic relationships between these bacteria.     

P2X<Subscript>2</Subscript> and P2X<Subscript>3</Subscript> purinoceptors in the rat enteric nervous system

Adenosine 5-triphosphate receptors are known to be involved in fast excitatory postsynaptic currents in myenteric neurons of the digestive tract. In the present study, the distribution of P2X₂ and P2X₃ receptor mRNA was examined by in situ hybridisation while P2X₂ and P2X₃ receptor protein was localised by immunohistochemical methods. In addition, P2X₂ and P2X₃ receptors were colocalised with calbindin and calretinin in the myenteric and submucosal plexus. P2X₂- and P2X₃-immunoreactive neurons were found in the myenteric and submucosal plexuses throughout the entire length of the rat digestive tract from the stomach to the colon. Approximately 60%, 70% and 50% of the ganglion cells in the myenteric plexus of the gastric corpus, ileum and distal colon, and 56% and 45% in the submucosal plexus of the ileum and distal colon, respectively, showed positive immunoreactivity to the P2X₂ receptor. Approximately 10%, 2% and 15% of the ganglion cells in the myenteric plexus of the gastric corpus, ileum and distal colon, and 62% and 40% in the submucosal plexus of the ileum and distal colon, respectively, showed positive immunoreactivity to the P2X₃ receptor. Double-labelling studies showed that about 10–25% of the neurons with P2X₂ immunoreactivity in myenteric plexus and 30–50% in the submucosal plexus were found to express calbindin or calretinin. About 80% of the neurons with P2X₃ receptor immunoreactivity in the myenteric plexus and about 40% in the submucosal plexus expressed calretinin. Approximately 30–75% of the neurons with P2X₃ receptor immunoreactivity in the submucosal plexus expressed calbindin, while none of them were found to express calbindin in the myenteric plexus.     

Replacing dietary glucose with fructose increases ChREBP activity and SREBP-1 protein in rat liver nucleus

Diets high in fructose cause hypertriglyceridemia and insulin resistance in part due to simultaneous induction of gluconeogenic and lipogenic genes in liver. We investigated the mechanism underlying the unique pattern of gene induction by dietary fructose. Male Sprague-Dawley rats (n = 6 per group) were meal-fed (4 h/d) either 63% (w/w) glucose or 63% fructose diet. After two weeks, animals were killed at the end of the last meal. Nuclear SREBP-1 was 2.2 times higher in fructose-fed rats than glucose-fed rats. Nuclear FoxO1 was elevated 1.7 times in fructose group, but did not reach significance (P = 0.08). Unexpectedly, no difference was observed in nuclear ChREBP between two groups. However, ChREBP DNA binding was 3.9× higher in fructose-fed animals without an increase in xylulose-5-phospate, a proposed ChREBP activator. In conclusion, the gene induction by dietary fructose is likely to be mediated in part by simultaneously increased ChREBP activity, SREBP-1 and possibly FoxO1 protein in nucleus.