Bacteriorhodopsin in model membranes. A new component of the displacement photocurrent in the microsecond time scale.

A quasi-short-circuit (tunable voltage clamp) measurement method with microsecond time resolution was applied to a bacteriorhodopsin model membrane formed by a novel interfacial technique. A new component (B1) of the displacement photocurrent was recorded: it has no detectable latency at an instrumental time constant of 1.5 museconds, and persists at 5 degrees C. In addition, a slower component (B2) of opposite polarity inhibited by low temperature (5 degrees C) and low pH (pH = 3.0) was recorded. The technique is very sensitive for the study of fast capacitative photoresponses in model membranes, and allows the detection of charge displacements in bacteriorhodopsin associated with distinct stages of the photochemical transformation.     

Effects of 2-deoxy-d-glucose,amiloride, vasopressin,and ouabain on active conductance andE Na in the toad bladder

Summary The effects of various agents on active sodium transport were studied in the toad bladder in terms of the equivalent circuit comprising an active conductanceK ^a, an electromotive forceE _Na, and a parallel passive conductanceK ^p. For agents which affectK ^a, but notE _Na orK ^p, the inverse slope of the plot of total conductance against short-circuit currentI ₀ evaluatesE _Na, and the intercept representsK ^p. Studies employing 5×10^–7 m amiloride to depressK ^a indicate a changingE _Na, invalidating the use of the slope technique with this agent. An alternative suitable technique employs 10^–5 m amiloride, which reducesI ₀ reversibly to near zero without effect onK ^p. Despite curvilinearity of the -I₀ plot under these conditions,K ^p may therefore be estimated fairly precisely from the residual conductance. It then becomes possible to follow the dynamic behavior ofK ^a andE _Na (in the absence of 10^–5 m amiloride) by frequent measurements of andI ₀, utilizing the relationshipsK ^a=K-K ^p, andK _Na=I _O/(K-K ^p). 2-deoxy-d-glucose (7.5×10^–3 m) depressedK ^a without affectingE _Na. Amiloride (5×10^–7 m) depressedK ^a and enhancedE _Na. Vasopressin (100 mU/ml) enhancedK ^a markedly and depressedE _Na slightly. Ouabain (10^–4 m) depressed bothK ^a andE _Na. All of the above effects were noted promptly;K ^p was unaffected. The electromotive force of Na transportE _Na appears not to be a pure energetic parameter, but to reflect kinetic factors as well, in accordance with thermodynamic considerations.     

The influence of insulin on glucose permeability and metabolism of human granulocytes.

Viable human polymorphonuclear leukocytes isolated from peripheral blood were incubated for 1 h at 37 degrees C with variable concentrations of insulin in a saline medium buffered at pH 7.4. The hormone increased glucose consumption by about 40% without influencing the permeability of the membranes to glucose, whose uptake followed a passive diffusion process. The measurement of intermediates localized activation of glycolysis by insulin, down to 0.36 nM, at the phosphofructokinase step. However, the spectrophotometric measurement showed no activation of phosphofructokinase after preincubation with insulin of either intact granulocytes or crude or ultracentrifuged homogenates. The level of cyclic AMP, which is known to activate phosphofructokinase, was not modified by insulin; cyclic GMP did not activate the enzyme in the granulocyte extracts: neither of the two nucleotides can therefore be considered as a direct messenger of the action of insulin on phosphofructokinase. An important fraction of the extra glucose consumed under the influence of insulin was recovered as neither glycogen nor lactate, nor was it oxidized in the Krebs cycle. It might be assumed to have been converted into glycerolipids. However, insulin produced no detectable accumulation of triglycerides and activated neither the pentose phosphate pathway nor oxidative decarboxylation of pyruvate. The fate of the extra glucose consumed under the influence of insulin therefore remains questionable.     

Different timing of increases in activities of four X-chromosome linked enzymes during the cell cycle of synchronized human lymphoblasts

A permanent human lymphoblast culture was synchronized with repetitive thymidine blocks, and the changes in the levels of activity of four X-chromosome-linked enzymes were followed during the cell cycle. The four enzymes studied were phosphoglycerate kinase (PGK), α-galactosidase (α-Gal), hypoxanthine-guanine phosphoribosyltransferase (HGPRT), and glucose-6-phosphate dehydrogenase (G6PD). The levels of PGK and α-Gal activities increased simultaneously in G1 and in S, while HGPRT and G6PD increased close together in middle and late S. Therefore, different control mechanisms may be involved in the increases of the activities of these two sets of enzymes.     

Changes in active transport, intracellular adenosine 5'-triphosphate levels, macromolecular syntheses, and glycolysis in an energy-uncoupled mutant of Escherichia coli.

The temperature-sensitive Escherichia coli mutant ecfts metC (Lieberman and Hong, 1974), previously shown to be defective in the coupling of metabolic energy to active transport, is also altered in a wide variety of cellular activities at the nonpermissive temperature. These alterations include a lowering of intracellular adenosine 5'-triphosphate levels, an alteration of glucose metabolism such that large quantities of pyruvate and dihydroxyacetone phosphate are excreted into the medium, excretion of accumulated potassium ions, and a cessation of deoxyribonucleic acid, ribonucleic acid, and phospholipid synthesis. Since these effects closely mimic the action of colicins E1 and K on E. coli cells, the possibility that the ecf gene product is the primary biochemical target for these colicins is discussed.     

Malocclusion of the premolar and molar teeth in the guinea pig.

Malocclusions of the premolar and molar teeth of inbred Strain 2/N and Strain 13/N and outbred Dunkin-Harley guinea pigs were examined. A higher incidence of malocclusion observed in the inbred strains suggested a genetic basis for the disorder.     

Constrained evolution of overlapping genes in viral host adaptation: Acquisition of glycosylation motifs in hepadnaviral precore/core genes

Hepadnaviruses use extensively overlapping genes to expand their coding capacity, especially the precore/core genes encode the precore and core proteins with mostly identical sequences but distinct functions. The precore protein of the woodchuck hepatitis virus (WHV) is N-glycosylated, in contrast to the precore of the human hepatitis B virus (HBV) that lacks N-glycosylation. To explore the roles of the N-linked glycosylation sites in precore and core functions, we substituted T77 and T92 in the WHV precore/core N-glycosylation motifs (⁷⁵NIT⁷⁷ and ⁹⁰NDT⁹²) with the corresponding HBV residues (E77 and N92) to eliminate the sequons. Conversely, these N-glycosylation sequons were introduced into the HBV precore/core gene by E77T and N92T substitutions. We found that N-glycosylation increased the levels of secreted precore gene products from both HBV and WHV. However, the HBV core (HBc) protein carrying the E77T substitution was defective in supporting virion secretion, and during infection, the HBc E77T and N92T substitutions impaired the formation of the covalently closed circular DNA (cccDNA), the critical viral DNA molecule responsible for establishing and maintaining infection. In cross-species complementation assays, both HBc and WHV core (WHc) proteins supported all steps of intracellular replication of the heterologous virus while WHc, with or without the N-glycosylation sequons, failed to interact with HBV envelope proteins for virion secretion. Interestingly, WHc supported more efficiently intracellular cccDNA amplification than HBc in the context of either HBV or WHV. These findings reveal novel determinants of precore secretion and core functions and illustrate strong constraints during viral host adaptation resulting from their compact genome and extensive use of overlapping genes.     

西藏东南部色季拉山主要类型森林叶片和枯落物养分含量特征

为阐明不同生长年限森林叶片和不同分解程度枯落物养分含量特征,为植物-土壤养分循环研究提供科学依据。以藏东南色季拉山几种典型森林植被(雪山杜鹃(Rhododendron aganniphum)、海拔4000 m和3900 m区域急尖长苞冷杉(Abies georgei var. smithii)、川滇高山栎(Quercus aquifolioides))为研究对象,分析了1年生和2年生植物叶片及不同分解程度枯落物有机碳(OC)、全氮(TN)、全磷(TP)和全钾(TK)含量。结果表明:色季拉山森林叶片和枯落物OC含量表现为2年生叶片1年生叶片未分解枯落物(ND)半分解枯落物(SD)完全分解枯落物(CD),即老叶片以C积累为主,而枯落物OC含量随分解程度的增加而下降,叶片OC平均含量(68.5%)显著高于中国平均水平(45.5%);叶片N、P、K含量表现为1年生2年生,即新叶以N、P、K等营养物质的吸收积累为主。枯落物TN含量低于中国森林的平均水平(12.03 g/kg),而TP含量显著高于中国森林平均水平(0.74 g/kg),枯落物TN和TP以SD最高,即分解初期表现为净固定,而后期则呈净释放,TK含量随分解程度的增加而增加,表现为K的净固定;叶片C∶N,C∶P和C∶K表现为2年生1年生,枯落物C∶N,C∶P和C∶K随着分解程度的增加而显著降低;叶片N∶P处于较低水平(6.08),显著低于全球平均水平(16.0),表现出明显的N限制营养型;研究结果为科学阐明藏东南森林生态系统植被-土壤养分循环研究提供了数据支撑。     

响应面模型分析高温蛋白酶菌株增殖和产酶关系

【目的】【方法】采用Box-Behnken法设计三因素三水平响应面实验,对比分析了高温蛋白酶高产菌株枯草芽孢杆菌BY25发酵过程中细菌生长与产酶之间的关系。【结果】实验表明,细菌生长与产酶的关系在各因素交互影响下变化显著。在中低水平的有机氮源添加条件下,细菌的大量繁殖显著抑制了蛋白酶产量,而在高水平的有机氮源添加条件下,细菌量的增长对产酶的作用由抑制转为促进。【结论】该结果表明,产酶诱导物有机氮源豆制品废渣的添加很可能显著提高了枯草芽孢杆菌群体中产酶菌的比率,而在没有合适诱导物的情况下通过增加细菌量并不能有效提高蛋白酶产量。