红光熊蜂线粒体CoⅠ基因的克隆与特性分析

通过对NCBI数据库的搜查,获得熊蜂CoI基因部分序列;设计引物,采用RT-PCR和RACE技术克隆了红光熊蜂CoI基因cDNA全长序列.采用DNAMAN 8.0,MEGA 7.0,Protparam和Automatic Modeling Mode等软件对该cDNA序列、基本理化性质、3-D结构、跨膜结构域及其系统进化关系进行了分析.结果表明:红光熊蜂CoI全长cDNA序列含1050 bp,编码349个氨基酸,对应的蛋白质相对分子质量为38350,预测其分子式为C1789 H2724 N422 O463 S25,理论等电点(pI)5.79,偏酸性,半衰期为30 h;蛋白质平均亲水系数0.706,不稳定性指数39.98,脂溶性指数110.92.该蛋白质具8个α-螺旋、8个跨膜结构域,为稳定的疏水性蛋白质.系统发育显示红光熊蜂Co I氨基酸序列与同属地熊蜂的进化关系较近,表明红光熊蜂线粒体Co I基因序列保守性较高.     

鞘氨醇单胞菌中威兰胶合成关键基因welE和welC的生物信息学分析

为探究威兰胶合成途径中关键基因welE和welC的结构特性及功能,运用生物信息学手段分析了welE和welC基因的序列信息,预测了其编码蛋白的理化性质、跨膜区、信号肽、磷酸化修饰、结构域及高级结构等.研究结果表明,WelE蛋白是由234个氨基酸组成的稳定亲水性蛋白质,无跨膜结构和信号肽,存在20个磷酸化位点,含有一个BY-kinase结构域,说明WelE可能参与威兰胶的生产与转运,WelE二级结构主要由α-螺旋和无规则卷曲组成,三级结构呈"圆环状";WelC蛋白是由449个氨基酸组成的不稳定亲水性跨膜蛋白质,含有2个跨膜螺旋、无信号肽,存在39个磷酸化位点,含有一个Wzz结构域,这表明WelC可能与威兰胶链长调节有关,WelC二级结构以α-螺旋和无规则卷曲为主,三级结构呈"条状",这些结果为解析welE和welC基因的功能以及建立威兰胶代谢调控方法奠定了理论基础.     

梅花鹿FGF10基因的生物信息学分析

应用生物信息学的方法对梅花鹿FGF10基因的核苷酸和氨基酸序列进行了初步的生物信息学分析,包括理化性质分析、信号肽和跨膜结构域分析、磷酸化位点和疏水性分析、蛋白质二级结构分析、功能结构域分析以及系统进化分析.结果表明:梅花鹿FGF10基因编码213个氨基酸,蛋白相对分子量为23.84kD,为碱性不稳定蛋白;存在信号肽和跨膜结构域;共有26个磷酸化位点;二级结构主要由α螺旋、β转角、延伸链和随机卷曲组成.具有FGF典型的FGF结构域.系统进化分析显示,梅花鹿FGF10与哺乳动物FGF10相似性较高,并且与牛、羊在亲缘关系上最相近.为梅花鹿FGF10基因的结构和功能的进一步研究打下了坚实的理论基础.     

甜瓜持绿蛋白的生物信息学分析

采用生物信息学方法,对甜瓜持绿蛋白(stay-green protein,SGR)理化性质、信号肽、前导肽、功能型位点、疏水性和亲水性、跨膜结构域、蛋白质二级结构和三级结构进行预测和分析,并构建系统进化树.结果表明:甜瓜含有4个持绿蛋白家族成员,进化分析发现有2个同源基因对,同源的基因对之间功能上完全冗余,4个蛋白都不具有信号肽和跨膜结构域,属于非分泌型蛋白,推测SGR可能在细胞质基质中发挥功能.采用同源建模法预测其三级结构,并通过拉氏构象图分析准确性和合理性,证明了同源建模的结果相对可靠性.     

中华鳖tyrp-1基因的生物信息学分析

为阐明中华鳖酪氨酸酶相关蛋白1(TYRP-1)基因tyrp-1及蛋白质结构与性质,利用相关软件、数据库和方法对其进行了生物信息学分析.结果显示,基因转录本全长为2 829bp,编码535个aa.预测TYRP-1蛋白分子质量为60.16ku,理论等电点为5.35,为不稳定蛋白;N端含有信号肽,与信号识别蛋白结合后,促进其穿越内质网膜进入内质网腔;C端含有跨膜结构域,锚定于细胞膜上,发挥重要的生物学功能;TYRP-1蛋白含有2个铜离子结合结构域,与DCT蛋白有明显的相互作用.     

Trichoderma Reesei菌生物降解酶CIP 1和CIP 2的生物信息学分析

运用生物信息学方法,对已报道的Trichoderma reesei QM6a菌株生物降解酶CIP 1和CIP 2基因进行了生物信息学分析,以明确里氏木霉生物降解酶CIP 1和CIP 2的理化性质、二级结构、信号肽、定位、跨膜结构、磷酸化位点及进化关系.结果表明：CIP 1和CIP 2均属于稳定蛋白,含有大量无规则卷曲;信号肽剪切位点分别在19~20和17~18位的氨基酸之间;两者无螺旋卷曲结构,无跨膜结构域,并定位在分泌途径信号肽（SP）上.     

东亚飞蝗气味结合蛋白Ⅰ的生物信息学分析

利用生物信息学方法分析了东亚飞蝗(Locusta migratoria manilensis)OBP1的核苷酸与氨基酸序列(GenBank登录号FJ215322.1/ACI30696.1),并对其组成成分、疏水/亲水区域、信号肽、跨膜结构域、蛋白质二级结构,以及分子进化关系等进行了预测与推断.结果显示,该蛋白由148个氨基酸组成,预测相对分子质量为16 122.6,理论等电点(pI)为4.99,分子式为C696H1128N184O223S15;含有蛋白激酶C磷酸化位点和酪氨酸激酶Ⅱ磷酸化位点等;具有标准的昆虫信息素/气味结合蛋白结构域.     

γ-聚谷氨酸合成酶系PgsBCA结构的生物信息学分析

利用ProtParam、TopPred、PredictProtein、PSORT-B Prediction、SWISS-MODEL等软件分别分析蛋白质的理化性质、跨膜区、二级结构、亚细胞定位、三维结构.结果显示:PgsB是亲水不稳定蛋白,通过豆蔻酰锚钩锚定于质膜上,催化作用需与ATP结合提供能量;PgsC是疏水稳定蛋白,通过4个跨膜区和多个豆蔻酰锚钩定位于质膜,具有酰胺化位点;PgsA是亲水稳定蛋白,通过N端一个跨膜区和豆蔻酰锚钩结合于质膜,具有多种磷酸化位点.说明γ-聚谷氨酸(Polyγ-glutamic acid,γ-PGA)合成酶系3个组分蛋白形成复合物定位于质膜上,其中PgsB在胞内催化γ-PGA合成,PgsC固定于质膜,连接PgsB和PgsA组分,PgsA在胞外负责γ-PGA的运输.通过对γ-PGA合成酶系各组分蛋白结构的分析,为日后在谷氨酸高产菌株中的表达奠定了基础.     

梅花鹿抗病毒蛋白的生物信息学分析

通过在线生物软件分析梅花鹿四种抗病毒蛋白A3Z2、BST-2A、BST-2B和SAMHD1蛋白的生物信息学特性。从转录组中获得四种蛋白的基因序列,应用Prot Param、Protscale、SOPMA、TMHMM、Target P、Signal P、Motif Scan、Interproscan以及BLAST等在线软件分析蛋白的理化性质、二级结构、穿膜域、结构域等等。首次获得了四种抗病毒因子的基因和蛋白序列,梅花鹿A3Z2基因编码393个氨基酸,相对分子质量为47.59 k Da,碱性,不稳定性亲水蛋白,无信号肽和跨膜结构域,胞内定位,具有糖基化和磷酸化位点,两个胞嘧啶脱氨酶结构域。梅花鹿BST-2A和2B基因编码159个氨基酸,相对分子质量为17.69 k Da和18.12 k Da,酸性,不稳定性亲水蛋白,无信号肽,BST-2A有两个跨膜结构域,BST-2B有一个跨膜结构域,膜定位,具有糖基化和磷酸化位点。梅花鹿SAMHD1基因编码613个氨基酸,相对分子质量为70.51 k Da,碱性,不稳定性亲水蛋白,无信号肽和跨膜结构域,胞内定位,具有磷酸化位点,d NTP磷酸水解酶活性结构域。梅花鹿A3Z2、BST-2A、BST-2B和SAMHD1蛋白具有潜在的抗病毒能力。     

Requirement of the RNA helicase-like protein PRP22 for release of messenger RNA from spliceosomes

The product of the yeast PRP22 gene acts late in the splicing of yeast pre-messenger RNA, mediating the release of the spliced mRNA from the spliceosome. The predicted PRP22 protein sequence shares extensive homology with that of PRP2 and PRP16 proteins, which are also involved in nuclear pre-mRNA splicing. The homologous region contains sequence elements characteristic of several demonstrated or putative ATP-dependent RNA helicases. A putative RNA-binding motif originally identified in bacterial ribosomal protein S1 and Escherichia coli polynucleotide phosphorylase has also been found in PRP22.     

拟南芥类结瘤素基因At1g01070的生物信息学分析

类结瘤素基因在非结瘤植物生长发育中承担着重要的功能,但仅少数基因的功能被鉴定.本研究对拟南芥类结瘤素MtN21(Medicago truncatula NODULIN 21)家族成员At1g01070进行了生物信息学分析及亚细胞定位验证.结果表明,At1g01070有两种可变剪切体,编码两种蛋白At1g01070.1和At1g01070.2,前者比后者在N端多47个氨基酸,它们的分子量分别为40KD和35KD,等电点分别为9.2和8.9;均含有2个MtN21蛋白的特征结构域EamA(药物/代谢物转运结构域),At1g01070.1比At1g01070.2多一个PLN00411结构域,且其靠近N端的EamA结构域较后者长;三级结构均由7个α-螺旋和一些不规则卷曲组成,分别含有10个和8个疏水跨膜区,均含13个磷酸化修饰位点,亚细胞定位预测主要定位在质膜;进化上,与琴叶拟南芥(Arabidopsis lyrata)亲缘关系最近;亚细胞定位结果显示At1g01070.1定位在质膜,与预测结果一致.推测At1g01070在植物体内可能参与物质运输.     

Structural basis for recognition of the tra mRNA precursor by the Sex-lethal protein

The Sex-lethal (Sxl) protein of Drosophila melanogaster regulates alternative splicing of the transformer (tra) messenger RNA precursor by binding to the tra polypyrimidine tract during the sex-determination process. The crystal structure has now been determined at 2.6 A resolution of the complex formed between two tandemly arranged RNA-binding domains of the Sxl protein and a 12-nucleotide, single-stranded RNA derived from the tra polypyrimidine tract. The two RNA-binding domains have their beta-sheet platforms facing each other to form a V-shaped cleft. The RNA is characteristically extended and bound in this cleft, where the UGUUUUUUU sequence is specifically recognized by the protein. This structure offers the first insight, to our knowledge, into how a protein binds specifically to a cognate RNA without any intramolecular base-pairing.     

Embryonic MAP2 lacks the cross-linking sidearm sequences and dendritic targeting signal of adult MAP2

The most prominent microtubule-associated protein of the neuronal cytoskeleton is MAP2. In the brain it exists as a pair of high-molecular weight proteins, MAP2a and MAP2b, and a smaller form, MAP2c, which is particularly abundant in the developing brain. High-molecular weight MAP2 is expressed in dendrites, where its messenger RNA is also located, but is not found in axons; it has been shown to be present in fine filaments that crosslink dendritic microtubules. This correlates with the primary structure of high-molecular weight MAP2, which consists of a short carboxy-terminal tubulin-binding domain and a long amino-terminal arm, which forms a filamentous sidearm on reconstituted microtubules. Here we report that the high- and low-molecular weight forms of MAP2 are generated by alternative splicing and share the entire C-terminal tubulin-binding domain as well as a short N-terminal sequence. In contrast to high molecular weight MAP2, embryonic brain MAP2c lacks 1,342 amino acids from the filamentous sidearm domain. Furthermore, the mRNA for low molecular weight MAP2c is not present in dendrites, indicating that the dendritic targeting signal is specific for the high-molecular weight form.     

The human RNA kinase hClp1 is active on 3' transfer RNA exons and short interfering RNAs

RNA interference allows the analysis of gene function by introducing synthetic, short interfering RNAs (siRNAs) into cells. In contrast to siRNA and microRNA duplexes generated endogenously by the RNaseIII endonuclease Dicer, synthetic siRNAs display a 5' OH group. However, to become incorporated into the RNA-induced silencing complex (RISC) and mediate target RNA cleavage, the guide strand of an siRNA needs to display a phosphate group at the 5' end. The identity of the responsible kinase has so far remained elusive. Monitoring siRNA phosphorylation, we applied a chromatographic approach that resulted in the identification of the protein hClp1 (human Clp1), a known component of both transfer RNA splicing and messenger RNA 3'-end formation machineries. Here we report that the kinase hClp1 phosphorylates and licenses synthetic siRNAs to become assembled into RISC for subsequent target RNA cleavage. More importantly, we reveal the physiological role of hClp1 as the RNA kinase that phosphorylates the 5' end of the 3' exon during human tRNA splicing, allowing the subsequent ligation of both exon halves by an unknown tRNA ligase. The investigation of this novel enzymatic activity of hClp1 in the context of mRNA 3'-end formation, where no RNA phosphorylation event has hitherto been predicted, remains a challenge for the future.