Interaction of preS domains of hepatitis B virus with phospholipid vesicles

The role of preS domains of the hepatitis B virus (HBV) envelope proteins in the first steps of viral infection has been restricted to their implication in virus attachment to a putative hepatocyte receptor. In order to explore a fusion activity in these regions, we used recombinant preS domains to characterize their interaction with liposomes. Binding experiments carried out with NBD-labeled proteins indicated that preS were able to interact in a monomeric way with acidic phospholipid vesicles, being the partition coefficient similar to that described for peptides which can insert deeply into bilayers. Fluorescence depolarization of DPH-labeled vesicles confirmed the specificity for negative charged phospholipids. Upon interaction the proteins induced aggregation, lipid mixing and release of internal contents of acidic vesicles at both acid and neutral pH in a concentration-dependent manner. Taken together, all these data indicate that preS domains are able to insert into the hydrophobic core of the bilayer. Moreover, the insertion resulted in a protein conformational change which increased the helical content. Therefore all these results suggest that, besides their participation in the recognition of a cellular receptor, the preS domains could be involved in the fusion mechanism of HBV with the plasma membrane of target cells.     

The N-terminal domain of the replication initiator protein RepE is a dimerization domain forming a stable dimer

The initiator protein RepE of the mini-F plasmid in Escherichia coli plays an essential role in DNA replication, which is regulated by the molecular chaperone-dependent oligomeric state (monomer or dimer). Crosslinking, ultracentrifugation, and gel filtration analyses showed that the solely expressed N-terminal domain (residues 1-144 or 1-152) exists in the dimeric state as in the wild-type RepE protein. This result indicates that the N-terminal domain functions as a dimerization domain of RepE and might be important for the interaction with the molecular chaperones. The N-terminal domain dimer has been crystallized in order to obtain structural insight into the regulation of the monomer/dimer conversion of RepE.     

Loop Domain Peptides from the SOS ras-Guanine Nucleotide Exchange Protein, Identified from Molecular Dynamics Calculations, Strongly Inhibit ras Signaling

In the accompanying paper, we found, using molecular dynamics calculations, four domains of the ras-specific SOS guanine nucleotide exchange protein (residues 589-601, 654-675, 746-761, and 980-989) that differ markedly in conformation when SOS is complexed with either oncogenic (Val 12-) ras-p21 or wild-type ras-p21. Three of these domains contain three crystallographically undefined loops that we modeled in these calculations, and one is a newly identified non-loop domain containing SOS residues 980-989. We have now synthesized peptides corresponding to these four domains and find that all of them block Val 12-ras-p21-induced oocyte maturation. All of them also block insulin-induced oocyte maturation, but two of these peptides, corresponding to SOS residues 589-601 and 980-989, block oncogenic ras to a significantly greater extent. These results suggest that SOS contains domains, including the three loop domains, that are important for ras signaling and that several of these domains can activate different pathways specific to oncogenic or wild-type ras-p21.     

The osmotic characteristics of human fetal liver-derived hematopoietic stem cell candidates

Hematopoietic stem cells derived from fetal liver have promising therapeutic potential for allotransplantation but require a specific protocol to minimize the damage produced by cryopreservation procedures. In this study, a fundamental approach was applied for designing a cell preservation protocol. To this end, the biophysical characteristics that describe the osmotic reaction of CD34(+)CD38(-) human fetal liver stem cell candidates were studied using fluorescent microscopy. The osmotically inactive volume of the stem cell candidates was determined as 48% of the isotonic volume. The permeability coefficients for water and Me(2)SO were determined at T = +22 degree C: L(p) = 0.27 +/- 0.03 microm x min(-1)atm(-1), P(Me(2)SO)) = 2.09 +/- 0.85 x 10 (-4) cm x min(-1), sigma (Me(2)SO)) = 0.63 +/- 0.03 and at T = +12 degree C: L(p) = 0.15 +/-0.02 microm x min(-1)atm(-1), P(Me(2)SO)) = 6.44 +/-1.42 x 10 (-5) cm x min(-1), sigma (Me(2)SO)) = 0.46 +/- 0.05. The results obtained suggest that post-hypertonic and hypotonic stress are the possible reasons for damage to a CD34(+)CD38(-) cell during the cryopreservation procedure.     

Structural characterization of the N-terminal mineral modification domains from the molluscan crystal-modulating biomineralization proteins, AP7 and AP24

The AP7 and AP24 proteins represent a class of mineral-interaction polypeptides that are found in the aragonite-containing nacre layer of mollusk shell (H. rufescens). These proteins have been shown to preferentially interfere with calcium carbonate mineral growth in vitro. It is believed that both proteins play an important role in aragonite polymorph selection in the mollusk shell. Previously, we demonstrated the 1-30 amino acid (AA) N-terminal sequences of AP7 and AP24 represent mineral interaction/modification domains in both proteins, as evidenced by their ability to frustrate calcium carbonate crystal growth at step edge regions. In this present report, using free N-terminal, C(alpha)-amide "capped" synthetic polypeptides representing the 1-30 AA regions of AP7 (AP7-1 polypeptide) and AP24 (AP24-1 polypeptide) and NMR spectroscopy, we confirm that both N-terminal sequences possess putative Ca (II) interaction polyanionic sequence regions (2 x -DD- in AP7-1, -DDDED- in AP24-1) that are random coil-like in structure. However, with regard to the remaining sequences regions, each polypeptide features unique structural differences. AP7-1 possesses an extended beta-strand or polyproline type II-like structure within the A11-M10, S12-V13, and S28-I27 sequence regions, with the remaining sequence regions adopting a random-coil-like structure, a trait common to other polyelectrolyte mineral-associated polypeptide sequences. Conversely, AP24-1 possesses random coil-like structure within A1-S9 and Q14-N16 sequence regions, and evidence for turn-like, bend, or loop conformation within the G10-N13, Q17-N24, and M29-F30 sequence regions, similar to the structures identified within the putative elastomeric proteins Lustrin A and sea urchin spicule matrix proteins. The similarities and differences in AP7 and AP24 N-terminal domain structure are discussed with regard to joint AP7-AP24 protein modification of calcium carbonate growth.     

Two Clip Domain Family of Serine Proteases and a Serine Protease Homolog from the Fall Webworm, Hyphantria cunea; cDNA Cloning and Characterization

Two types of serine proteases and a serine protease homologue cDNAs were isolated from Hyphantria cunea larvae induced immune response due to an injection of a microorganism through RT‐PCR and cDNA library screening, and their characteristics were examined. The isolated cDNAs are composed 2.1 kb, 2.2 kb, and 2.5 kb nucleotide each, which encoded 388, 390, 580 amino acid residues, and were designated as HcPE‐1, HcPE‐2 and HcPE‐3, respectively. They were revealed as serine proteases or a serine protease homologue with the clip domain through a database search. The deduced amino acid sequence comparison showed high homology of 72‐78% among them. Six Cys residues of the N‐terminal clip domain forming the disulfide bond, Cys residues of the catalytic domain, and Cys residues forming inter‐bridge between clip domain and catalytic domain were also well preserved. Three amino acid residues, His, Asp, and Ser, within the active site were perfectly conserved in HcPE‐2 and HcPE‐3, however, His was replaced with Gln¹⁷⁸ in HcPE‐1. The Arg residues (HcPE‐1, Arg¹³²; HcPE‐2, Arg¹³⁴; HcPE‐3, Arg³²⁵) known as the activation sites by proteolytic cleavage were preserved well in all three types of protein. In case of HcPE‐3, three continuous clip‐like domains existed in the N terminal. As the result of phylogenetic analysis, three clip domain family of protein from H. cunea make groups with arthropod proclotting enzyme precursor. Northern blot analysis showed all three genes were induced through an injection of Escherichia coli, but expression patterns were varied.     

剑阁县2001～2010 年法定传染病流行特征及防治对策分析

目的:通过分析10年法定传染病疲情的流行趋势和三间分布特征,为制定传染病预防控制策略和措施提供依据.方法:采用描述性流行病学方法分析疫情趋势和三间分布情况,数据资料用SPSS10.0和Excel 2003进行统计分析.结果:2001～2010年共报告乙、丙类传染病25种26 129例,年均发病率386.89/10万,年均死亡率0.15/10万,10年间报告法定传染病以血源及性传播传染病和呼吸道传染病为主,居第1位的是血源及性传播传染病,共报告5种12 453例,占53.03％;其次是呼吸道传染病,共报告5种9828例,占41.85％,近3年发病居于各类传染病首位;第三位的是肠道传染病,共报5种1149例,占4.89％.发病居前5位的传染为乙肝、肺结核、流行性腮腺炎、痢疾、麻疹,主要传染病以乙肝、肺结核为主,近年性传播疾病呈快速增长趋势.结论:血源及性传播传染病和呼吸道传染病是今后重点防控传染病.     

福建和浙江乡村人居林结构特征比较

对位于福建省和浙江省8个地区39个村庄的乡村人居林4种林型(庭院林、道路林、水岸林和游憩林)的树木结构特征进行了调查和分析.结果表明:两省的乡村人居林共有乔木61科130属201种,共有灌木34科52属76种;福建省乡村人居林中乔木种类较为丰富;浙江省乡村人居林中灌木种类较为丰富.4种林型中,庭院林树种数最多,道路林树种数最少;组成庭院林的乔木树种和单株数均最多,而组成道路林的乔木树种及单株数均最少;组成庭院林和游憩林的灌木树种和单株(丛)数均较多,组成水岸林的灌木树种及单株(丛)数最少.在福建省和浙江省乡村人居林中,常绿树种、乡土树种和2003年以前种植的树种株数较多,分别占两省乡村人居林树木总株数的71.05％、85.57％、61.20％和65.95％、76.02％、61.43％;栽植截干和管护截干株数均较少,但浙江省乡村人居林中栽植截干和管护截干的株数均多于福建省.福建省乡村人居林中乔木的胸径、树高、冠幅和枝下高的均值均明显大于浙江省,而两省灌木的地径、树高、冠幅和枝下高的均值差异较小.福建省的游憩林及浙江省的水岸林中乔木的胸径最大,而福建省的水岸林及浙江省的庭院林中灌木的地径最大;福建省乡村人居林中胸径10.0 cm以上的乔木所占比例高于浙江省,其中福建省的4种林型中胸径在30.0 cm以上的乔木树种的平均胸径、种类数及株数均高于浙江省.两省乡村人居林中频度最高的10个树种明显不同,仅香樟[ Cinnamomum camphora (L.)Presl]共存于两省所有林型中.     

基于编码序列、基因间序列和氨基酸序列构建的系统发生关系比较

以7种古菌、46种细菌和10种真核生物的基因组为样本,考虑碱基间的短程关联和长程关联作用,得到编码序列的密码对和基因间序列的三联体对中不同位点的二核苷酸频率,据此构建了基于编码序列和基因间序列的系统发生关系。无论是基于编码序列还是基因间序列对信息进行聚类,古菌或真核均被聚在一支上,表明聚类参数的选择是合适的;与基于氨基酸序列构建的系统发生关系进行两两比较,发现大部分硬壁菌的编码序列与基因间序列之间,以及编码序列与氨基酸序列之间的进化都存在较大差异。通过分析认为,只有综合考虑这三类序列的进化信息,才可能得到更自然的系统发生关系。     

NaCl和Na2CO3胁迫对桑树幼苗生长和光合特性的影响

以1年生“青龙桑”幼苗为试验材料,研究了中性盐(NaCl)和碱性盐(Na2CO3)胁迫下桑树幼苗的生长和叶片光合特性.结果表明： 盐胁迫明显降低了桑树幼苗的株高、叶片数、生物量和叶片的光合能力.随着Na+浓度的增加,桑树叶片的气孔导度、蒸腾速率、净光合速率、实际光化学效率、电子传递速率和光化学猝灭系数明显降低,过剩光能以非光化学猝灭形式耗散的比例增加,桑树叶片的光能转化效率和光合能力下降.在Na+浓度＜150 mmol·L-1时,桑树幼苗的光合能力和生长受到的抑制较小,通过增加根冠比进一步适应盐胁迫,但这种保护机制随着盐浓度的增加逐渐降低.在Na2CO3胁迫下,＞50 mmol·L-1Na+浓度对桑树的生长和光合能力表现出较强的抑制作用,并随Na+浓度的增加,抑制程度加大.在NaCl＜150 mmol·L-1时,桑树的光合能力主要依赖植株形态和光合代谢双重途径适应中性盐逆境,而在NaCl浓度＞150 mmol·L-1和碱性盐胁迫下,其主要依赖光合代谢来适应逆境.