Short-term succession of marine microbial fouling communities and the identification of primary and secondary colonizers

Microbial succession during the initial stages of marine biofouling has been rarely studied, especially in the Arabian Gulf. This study was undertaken to follow temporal shifts in biofouling communities in order to identify primary and secondary colonizers. Quantitative analysis revealed a significant increase in total biomass, coverage of macrofoulers, chlorophyll a concentrations, and bacterial counts with time. The relative abundance of the adnate diatoms increased with time, whereas it decreased in the case of the plocon diatoms. Non-metric multidimensional scaling (NMDS) ordination based on MiSeq data placed the bacterial communities in three distinct clusters, depending on the time of sampling. While the relative abundance of Alphaproteobacteria and Flavobacteriia decreased with time, suggesting their role as primary colonizers, the relative abundance of Actinobacteria and Planctomycetia increased with time, suggesting their role as secondary colonizers. Biofouling is a dynamic process that involves temporal quantitative and qualitative shifts in the micro- and macrofouling communities.     

Mechanistic Studies on Metal-pyrophosphate Hydrolysis. Structure of Tetraammine(chloroaquo)cobait(III) dichloride ([CO(NH3)4ClH20]2+•-2Cl−), an Acid Hydrolysis Product of Co(NH3)4HP2O7 and Co(NH3)4PO4

Abstract

The octahedral complex tetraammine(chloroaquo)cobalt(III) dichloride is shown to be the HCl hydrolysis product of both P¹,^2-bidentate tetraammine(pyrophosphato)cobalt(III) [CO(NH₃)₄HP₂0₇ or CoPP] and bidentate tetraammine(phosphato)cobalt(III) [Co(NH₃)₄P0₄or CoP]. The complex crystallizes in the orthorhombic space group Pna2¹ with cell dimensions α=13.033(2)Å, b=6.710(1) Å, and c=10.318(2)Å; the crystal structure was refined to a final disagreement index of 0.033. The average of the four Co-N distances is 1.944±6Å. The Co-Cl distance is 2.257(2)Å and the Co-O(W) distance is 1.971(4)Å. Both protons of the coordinated water molecule are engaged in strong hydrogen bonds to the two nonbonded chloride counterions with 0(W)-C1 distances of 3.087(6)Å and 3.123(6)Å. Each nonbonded chloride is engaged in seven hydrogen bonding interactions resulting from the high ratio of hydrogen bond donors to acceptors in the CoP structure. Cobalt bisphosphate (CoP₂) is the final enzyme hydrolysis product when CoPP is used as substrate in the yeast inorganic pyrophosphatase reaction. The bridge oxygen atom is the site of initial CoPP cleavage both, for HCl catalyzed hydrolysis as well as for enzyme catalyzed hydrolysis.     

Resveratrol Binding to Human Serum Albumin

Abstract

Resveratrol (Res), a polyphenolic compound found largely in the skin of red grape and wine, exhibits a wide range of pharmaceutical properties and plays a role in prevention of human cardiovascular diseases [Pendurthi et al., Arterioscler. Thromb. Vasc. Biol. 19, 419–426 (1999)]. It shows a strong affinity towards protein binding and used as inhibitor for cyclo- oxygenase and ribonuclease reductase. The aim of this study was to examine the interaction of resveratrol with human serum albumin (HSA) in aqueous solution at physiological conditions, using a constant protein concentration (0.3 mM) and various pigment contents μM to mM). FTIR, UV-Visible, CD, and fluorescence spectroscopic methods were used to determine the resveratrol binding mode, the binding constant and the effects of pigment complexation on protein secondary structure.

Structural analysis showed that resveratrol bind non-specifically (H-bonding) via polypeptide polar groups with overall binding constant of K_Res = 2.56× 10⁵ M^?1. The protein secondary structure, analysed by CD spectroscopy, showed no major alterations at low resveratrol concentrations (0.125 mM), whereas at high pigment content (1 mM), major increase of α-helix from 57% (free HSA) to 62% and a decrease of β-sheet from 10% (free HSA) to 7% occurred in the resveratrol-HSA complexes. The results indicate a partial stabilization of protein secondary structure at high resveratrol content.     

西花蓟马取食与机械损伤对菜豆叶片抗氧化系统的影响

本文研究了西花蓟马Frankliniella occidentalis(Pergande)取食和机械损伤诱导对菜豆抗氧化系统的影响,比较了不同损伤形式诱导的抗氧化酶活力和抗氧化物质含量的变化差异。结果表明,西花蓟马取食和机械损伤均可诱导菜豆叶片内过氧化物酶(POD)、超氧化物歧化酶(SOD)和过氧化氢酶(CAT)3种抗氧化酶活性有不同程度的升高,但两种处理诱导的抗氧化酶活性变化规律不完全相同,各种酶活性达到最高点时间不同,西花蓟马取食对抗氧化酶活力的诱导作用大于机械损伤的。两种处理诱导下的类胡萝卜素的含量变化不大,但不论西花蓟马取食还是机械损伤均导致菜豆叶片内类黄酮和总酚含量整体呈下降趋势,西花蓟马的取食诱导的下降幅度大于机械损伤的。因此,西花蓟马取食诱导明显高于机械损伤对菜豆抗氧化系统的影响。     

Ulex europaeus Agglutinin-I Binding to Dental Primary Afferent Projections in the Spinal Trigeminal Complex Combined with Double Immunolabeling of Substance P and GABA Elements Using Peroxidase and Colloidal Gold

Ulex europaeus agglutinin I (UEA-I) is a plant lectin with an affinity for L-fucosyl residues in the chains of lactoseries oligosaccharides associated with medium and smaller-diameter dorsal root ganglion neurons and their axonal processes. These enter Lissauer's tract and terminate within the superficial laminae of the spinal cord overlapping projections known to have a nociceptive function. This implies that the surface coatings of neuronal membranes may have a relationship with functional modalities. The present investigation further examined this concept by studying a neuronal projection with a nociceptive function to determine whether fucosyl-lactoseries residues were incorporated in its primary afferent terminals. Transganglionic transport of horseradish peroxidase (HRP) following injection into tooth pulp chambers was employed to demonstrate dental pulp terminals in the trigeminal spinal complex, while peroxidase and fluorescent tags were used concomitantly to stain for UEA-I. Double immunolabeling for substance P (SP) and γ-aminobutyric acid (GABA) using peroxidase and colloidal gold allowed a comparison of the distribution of a known excitatory nociceptive transmitter with that of UEA-I binding in specific subnuclei. Synaptic interrelationships between UEA-I positive dental pulp primary afferent inputs and specific inhibitory terminals were also examined.

SP immunoreactivity occurred in laminae I and outer lamina II (II_o) of subnucleus caudalis (Vc) and in the ventrolateral and lateral marginal region of the caudal half of subnucleus interpolaris (Vi), including the periobex area in which Vi is slightly overlapped on its lateral aspect by cellular elements of Vc. The adjacent interstitial nucleus (IN) also showed an intense immunoreactivity for this peptide antibody. UEA-I binding displayed a similar distribution pattern in both Vc and Vi, but extended into lamina II; and the superficial part of Lamina III in Vc. Dental pulp terminals were found to have a comparable distribution; however, many extended into the dorsal portion of the caudal half of Vi and the ventromedial quadrant of rostral Vi.

Electron-microscopic analysis showed that transganglionically labeled dental pulp terminals contained ovoid, complex membrane-bound vacuoles laden with transported HRP. The preterminal axon and synaptic membranes of those dental pulp terminals located in zones of Vc and Vi displaying an affinity for UEA-I were usually characterized by a patchy, electron-dense coating of the peroxidase tag. SP was demonstrated ultrastructurally with Protein-A colloidal gold (3-nm particles), whereas GABA immunoreactivity was revealed by the avidin—biotin—peroxidase method. This combined approach labeled a variety of simple axodendritic to large complex scalloped dental terminals which contained SP and were shown to have a UEA-I affinity. In addition, many of the larger terminals formed contacts with GABA-ergic dendrites and received inputs from GABA-ergic synapses. These complexes were most concentrated in lamina II_o of Vc and the ventrolateral zone of Vi. Many terminals in laminae II_i; and III with a UEA-I-positive surface coating failed to bind with the antiserum for SP, indicating that other transmitters may colocalize with UEA-I and suggesting that absolute correlations between specific oligosaccharide plasmalemmal coatings and functional modalities should be approached with caution. Further studies employing antisera to different transmitters are currently underway to better define the relationship between transmitter localization and anatomical substrates within this circuitry. These studies should eventually provide additional clues about relationships between functional properties and oligosaccharide coatings of primary afferent projections.     

Novel Key Metabolites Reveal Further Branching of the Roquefortine/Meleagrin Biosynthetic Pathway

Metabolic profiling and structural elucidation of novel secondary metabolites obtained from derived deletion strains of the filamentous fungus Penicillium chrysogenum were used to reassign various previously ascribed synthetase genes of the roquefortine/meleagrin pathway to their corresponding products. Next to the structural characterization of roquefortine F and neoxaline, which are for the first time reported for P. chrysogenum, we identified the novel metabolite roquefortine L, including its degradation products, harboring remarkable chemical structures. Their biosynthesis is discussed, questioning the exclusive role of glandicoline A as key intermediate in the pathway. The results reveal that further enzymes of this pathway are rather unspecific and catalyze more than one reaction, leading to excessive branching in the pathway with meleagrin and neoxaline as end products of two branches.     

The Strawberry Pathogenesis-related 10 (PR-10) Fra a Proteins Control Flavonoid Biosynthesis by Binding to Metabolic Intermediates

Pathogenesis-related 10 (PR-10) proteins are involved in many aspects of plant biology but their molecular function is still unclear. They are related by sequence and structural homology to mammalian lipid transport and plant abscisic acid receptor proteins and are predicted to have cavities for ligand binding. Recently, three new members of the PR-10 family, the Fra a proteins, have been identified in strawberry, where they are required for the activity of the flavonoid biosynthesis pathway, which is essential for the development of color and flavor in fruits. Here, we show that Fra a proteins bind natural flavonoids with different selectivity and affinities in the low μm range. The structural analysis of Fra a 1 E and a Fra a 3-catechin complex indicates that loops L3, L5, and L7 surrounding the ligand-binding cavity show significant flexibility in the apo forms but close over the ligand in the Fra a 3-catechin complex. Our findings provide mechanistic insight on the function of Fra a proteins and suggest that PR-10 proteins, which are widespread in plants, may play a role in the control of secondary metabolic pathways by binding to metabolic intermediates.     

Advanced glycation end products suppress osteoblastic differentiation of stromal cells by activating endoplasmic reticulum stress

Advanced glycation end products (AGEs) are involved in bone quality deterioration in diabetes mellitus. We previously showed that AGE2 or AGE3 inhibited osteoblastic differentiation and mineralization of mouse stromal ST2 cells, and also induced apoptosis and decreased cell growth. Although quality management for synthesized proteins in endoplasmic reticulum (ER) is crucial for the maturation of osteoblasts, the effects of AGEs on ER stress in osteoblast lineage are unknown. We thus examined roles of ER stress in AGE2- or AGE3-induced suppression of osteoblastogenesis of ST2 cells. An ER stress inducer, thapsigargin (TG), induced osteoblastic differentiation of ST2 cells by increasing the levels of Osterix, type 1 collagen (Col1), alkaline phosphatase (ALP) and osteocalcin (OCN) mRNA. AGE2 or AGE3 suppressed the levels of ER stress sensors such as IRE1α, ATF6 and OASIS, while they increased the levels of PERK and its downstream molecules, ATF4. A reduction in PERK level by siRNA did not affect the AGEs-induced suppression of the levels of Osterix, Col1 and OCN mRNA. In conclusion, AGEs inhibited the osteoblastic differentiation of stromal cells by suppressing ER stress sensors and accumulating abnormal proteins in the cells. This process might accelerate AGEs-induced suppression of bone formation found in diabetes mellitus.     

Sequence and hydropathy profile analysis of two classes of secondary transporters

A structural class in the MemGen classification of membrane proteins is a set of evolutionary related proteins sharing a similar global fold. A structural class contains both closely related pairs of proteins for which homology is clear from sequence comparison and very distantly related pairs, for which it is not possible to establish homology based on sequence similarity alone. In the latter case the evolutionary link is based on hydropathy profile analysis. Here, we use these evolutionary related sets of proteins to analyze the relationship between E-values in BLAST searches, sequence similarities in multiple sequence alignments and structural similarities in hydropathy profile analyses. Two structural classes of secondary transporters termed ST[3], which includes the Ion Transporter (IT) superfamily and ST[4], which includes the DAACS family (TC# 2.A.23) were extracted from the NCBI protein database. ST[3] contains 2051 unique sequences distributed over 32 families and 59 subfamilies. ST[4] is a smaller class containing 399 unique sequences distributed over 2 families and 7 subfamilies. One subfamily in ST[4] contains a new class of binding protein dependent secondary transporters. Comparison of the averaged hydropathy profiles of the subfamilies in ST[3] and ST[4] revealed that the two classes represent different folds. Divergence of the sequences in ST[4] is much smaller than observed in ST[3], suggesting different constraints on the proteins during evolution. Analysis of the correlation between the evolutionary relationship of pairs of proteins in a class and the BLAST E-value revealed that: (i) the BLAST algorithm is unable to pick up the majority of the links between proteins in structural class ST[3], (ii) ‘low complexity filtering’ and ‘composition based statistics’ improve the specificity, but strongly reduce the sensitivity of BLAST searches for distantly related proteins, indicating that these filters are too stringent for the proteins analyzed, and (iii) the E-value cut-off, which may be used to evaluate evolutionary significance of a hit in a BLAST search is very different for the two structural classes of membrane proteins.     

邻苯二甲酸对萝卜种子萌发、幼苗叶片膜脂过氧化及渗透调节物质的影响

华北地区是我国玉米的主产区,玉米秸秆还田不仅可有效改善土壤理化性状、提高土壤生物有效性,还会在腐解过程中释放出目前公认的化感物质——酚酸类物质,邻苯二甲酸是玉米秸秆腐解液中的主要酚酸类物质。从玉米秸秆还田过程中主要腐解产物(邻苯二甲酸)对蔬菜作物的化感效应角度进行了研究,为量化秸秆还田量及构建粮-菜轮作制度探寻化感效应依据。试验以萝卜为蔬菜材料,通过配置4个浓度(0.05、0.5、1.0、2.0 mmol/L)的邻苯二甲酸溶液,模拟玉米秸秆还田条件,以清水为对照,研究主要腐解产物邻苯二甲酸对萝卜种子萌发、幼苗生长、膜脂过氧化作用及渗透调节物质的影响。结果表明:(1)萝卜不同生育期对邻苯二甲酸化感效应的响应程度不同。在0.05-1 mmol/L浓度范围内,邻苯二甲酸处理促进了萝卜种子萌发,但随着处理浓度的增大,促进作用减弱;浓度达到2 mmol/L时对萝卜种子萌发具有抑制效果。(2)邻苯二甲酸0.05mmol/L处理,促进了萝卜幼苗干鲜物质积累,幼苗根系生长,其中根系长度和根系表面积分别比对照提高42.03%、38.36%,显著高于清水对照;植株体内超氧化物歧化酶(Superoxide dismutase, SOD)活性增大,过氧化物酶(Peroxidase, POD)、过氧化氢酶(Catalase, CAT)活性降低,膜脂过氧化产物丙二醛(Malonaldehyde, MDA)含量与对照无显著差异。(3)当邻苯二甲酸浓度超过0.5 mmol/L时,萝卜幼苗脂质过氧化伤害加剧,体内MDA含量急剧增加,代谢与生理功能出现紊乱,正常生长及干鲜物质积累受到显著抑制。邻苯二甲酸浓度达到2 mmol/L时,叶片数较对照降低了36.51%;根系长度、根系表面积及根尖数降幅分别为64.46%、40.20%、41.28%。(4)对于渗透调节物质的影响,邻苯二甲酸处理促进了萝卜幼苗叶片可溶性糖含量的增加,但随着处理浓度的升高其促进作用逐渐减弱;可溶性蛋白含量随着邻苯二甲酸处理浓度的升高表现出逐渐减少的趋势,分别较对照降低了12.82%、14.88%、21.58%、24.73%。因此,华北地区实施玉米-萝卜轮作模式,从化感效应角度研究玉米秸秆量化还田,应将土壤中邻苯二甲酸浓度控制在0.5 mmol/L范围以内,以防止邻苯二甲酸浓度过高对萝卜幼苗生长的抑制作用。