【目的】研究鸡gga-mir-1658前体区基因遗传变异/单倍型及其在品种间的分布,分析其对micro RNA二级茎环结构和靶基因选择的影响,旨在筛选其中具有潜在生物学功能的变异位点,为进一步揭示其对gga-mir-1658基因表达调控的影响及表型效应...【目的】研究鸡gga-mir-1658前体区基因遗传变异/单倍型及其在品种间的分布,分析其对micro RNA二级茎环结构和靶基因选择的影响,旨在筛选其中具有潜在生物学功能的变异位点,为进一步揭示其对gga-mir-1658基因表达调控的影响及表型效应奠定基础。【方法】根据鸡gga-mir-1658基因组序列(Gen Bank登录号:NR_035151.1)设计一对特异性引物,采用PCR产物直接测序的方法,对太行鸡(95只)、北京油鸡(83只)和来航鸡(42只)3个鸡种220只个体的gga-mir-1658基因前体区进行多态性检测。使用DNAman、MEGA和mfold软件进行pre-gga-mir-1658基因组序列的比对分析和二级结构模拟。通过SHEsis软件和Haploview软件进行配对连锁不平衡分析和单倍型分析。使用mi Randa软件对gga-mir-1658的靶基因及其复合物自由能变化进行预测分析。【结果】在pre-gga-mir-1658基因共发现6个变异位点,其中次要等位基因频率≥5%位点有g.28 C>G、g.31C>T、g.70 G>A和g.71 G>–,4个变异位点均定位于gga-mir-1658基因成熟体的种子区。遗传变异特征分析显示,g.70 G>A位点表现为低度多态(PIC<0.25),其余3个位点均表现为中度多态(0.25<PIC<0.50)。适合性检验表明,除了来航鸡的g.31 C>T、g.71 G>–位点、太行鸡的g.71 G>–位点以及北京油鸡的g.70 G>A位点之外,其他变异位点在各鸡种均处于哈代-温伯格平衡状态(P>0.05)。连锁不平衡和单倍型分析表明,各突变位点之间存在弱连锁平衡;从3个品种鸡中共检测到11种单倍型,其中H1(C C G–)和H11(G T G G)是群体的优势单倍型,频率均大于25%。生物信息学分析表明,种子区的突变可影响gga-mir-1658基因前体二级结构的空间构型和自由能,其中H6单倍型突变体的自由能最高(41.00 kcal·mol-1),H2和H5单倍型突变体的自由能最低(35.70 kcal·mol-1);群体的优势单倍型H1和H11突变体的自由能分别为-36.10和40.04 kcal·mol-1。gga-mir-1658基因不同单倍型成熟体种子区序列存在差异,在gga-mir-1658-5p存在"AUACCAU"、"AUACCAC"2种种子区序列,gga-mir-1658-3p共有"AACUCUG"、"AGCUGUG"、"AACUGUG"和"AGCUCUG"4种种子区序列。针对gga-mir-1658的预测靶基因的生物信息学分析显示,它们主要在基因表达调控、细胞凋亡、免疫系统的发育和B细胞激活等基本生物学过程中显著富集;此外,种子区变化可以影响gga-mir-1658成熟体对靶基因的选择。【结论】(1)gga-mir-1658基因种子区存在4个具有潜在生物学功能和表型效应的突变位点,可组成11种单倍型,其中H1(C C G–)和H11(G T G G)在北京油鸡、太行鸡和来航鸡为优势单倍型。(2)种子区突变影响gga-mir-1658基因前体二级结构的稳定性和靶基因的选择,可能是具有潜在表型效应的重要功能位点。展开更多
The secondary structure of different Iβ cellulose was analyzed by a molecular dynamics sim- ulation with MARTINI coarse-grained force field, where each chain of the cellulose includes 40 D-glucoses units. Calculation...The secondary structure of different Iβ cellulose was analyzed by a molecular dynamics sim- ulation with MARTINI coarse-grained force field, where each chain of the cellulose includes 40 D-glucoses units. Calculation gives a satisfied description about the secondary structure of the cellulose. As the chain number increasing, the cellulose becomes the form of a helix, with the diameter of screw growing and spiral rising. Interestingly, the celluloses with chain number N of 4, 6, 24 and 36 do show right-hand twisting. On the contrast, the celluloses with N of 8, 12, 16 chains are left-hand twisting. These simulations indicate that the cellulose with chain number larger than 36 will break down to two parts. Besides, the result indicates that 36-chains cellulose model is the most stable among all models. Furthermore, the Lennard-Jones potential determines the secondary structure. In addition, an equation was set up to analyze the twisting structure.展开更多
One of the major obstacles facing the field of structural biology in the post genomic era is the inherent difficulty of analyzing the structure of membrane proteins under native conditions. The method of choice for st...One of the major obstacles facing the field of structural biology in the post genomic era is the inherent difficulty of analyzing the structure of membrane proteins under native conditions. The method of choice for studying such proteins is FTIR spectroscopy. Following the outbreaking of the severe acute respiratory syndrome （SARS） virus, in 2003, extensive work has been directed at elucidating the structure of the E transmembrane proteins of the SARS coronavirus. In this study, the secondary structure of the transmembrane a-helical bundles was analysised using the biophysical method site specific infrared dichroism （SSID）. Sixteen amino acids were isotopically labeled with （~3C=180） at different positions of the primary structure of the synthesized E protein CoV. The secondary structure was studied using Attenuated Total Internal Reflection （ATR） FTIR spectroscopy. Based on our findings, the presence of two possible H-bonding interactions between the carbonyl oxygen of two residues 26 and 31 （Phe and Leu） respectively with water molecules which may be trapped within the helix structure were postulatesed. These interactions may cause a change in this structure.展开更多
The native protein structures in buffer solution are maintained by the electrostatic force as well as the hydrophobic force, salt ions play an important role in maintaining the protein native structures, and their eff...The native protein structures in buffer solution are maintained by the electrostatic force as well as the hydrophobic force, salt ions play an important role in maintaining the protein native structures, and their effect on the protein stability has attracted tremendous interests. Infrared spectroscopy has been generally used in molecular structure analysis due to its fingerprint resolution for different species including macromolecules as proteins. However spectral intensities have received much less attention than the vibrational frequencies. Here we report that the spectral intensities of protein amide I band, the finger prints for the protein secondary structures, are very sensitive to the local electric field known as Onsager reaction field caused by salt ions. IR absorbance thermal titrations have been conducted for a series of samples including simple water soluble amino acids, water soluble monomeric protein cytochrome c and dimeric protein DsbC and its single-site mutant G49R. We found that at lower temperature range （10-20℃）, there exists a thermal activated salting-in process, where the IR intensity increases with a rise in the temperature, corresponding to the ions binding of the hydrophobic surface of protein. This process is absent for the amino acids. When further raising the temperature, the IR intensity decreases, this is interpreted as the thermal activated breaking of the ion-protein surface binding. Applying Van＇t Hoff plot to the thermal titration curves, the thermodynamic parameters such as AH and AS for salting-in and ion unbinding processes can be derived for various protein secondary structural components, revealing quantitatively the extent of hydrophobic interaction as well as the strength of the ion-protein binding.展开更多
文摘【目的】研究鸡gga-mir-1658前体区基因遗传变异/单倍型及其在品种间的分布,分析其对micro RNA二级茎环结构和靶基因选择的影响,旨在筛选其中具有潜在生物学功能的变异位点,为进一步揭示其对gga-mir-1658基因表达调控的影响及表型效应奠定基础。【方法】根据鸡gga-mir-1658基因组序列(Gen Bank登录号:NR_035151.1)设计一对特异性引物,采用PCR产物直接测序的方法,对太行鸡(95只)、北京油鸡(83只)和来航鸡(42只)3个鸡种220只个体的gga-mir-1658基因前体区进行多态性检测。使用DNAman、MEGA和mfold软件进行pre-gga-mir-1658基因组序列的比对分析和二级结构模拟。通过SHEsis软件和Haploview软件进行配对连锁不平衡分析和单倍型分析。使用mi Randa软件对gga-mir-1658的靶基因及其复合物自由能变化进行预测分析。【结果】在pre-gga-mir-1658基因共发现6个变异位点,其中次要等位基因频率≥5%位点有g.28 C>G、g.31C>T、g.70 G>A和g.71 G>–,4个变异位点均定位于gga-mir-1658基因成熟体的种子区。遗传变异特征分析显示,g.70 G>A位点表现为低度多态(PIC<0.25),其余3个位点均表现为中度多态(0.25<PIC<0.50)。适合性检验表明,除了来航鸡的g.31 C>T、g.71 G>–位点、太行鸡的g.71 G>–位点以及北京油鸡的g.70 G>A位点之外,其他变异位点在各鸡种均处于哈代-温伯格平衡状态(P>0.05)。连锁不平衡和单倍型分析表明,各突变位点之间存在弱连锁平衡;从3个品种鸡中共检测到11种单倍型,其中H1(C C G–)和H11(G T G G)是群体的优势单倍型,频率均大于25%。生物信息学分析表明,种子区的突变可影响gga-mir-1658基因前体二级结构的空间构型和自由能,其中H6单倍型突变体的自由能最高(41.00 kcal·mol-1),H2和H5单倍型突变体的自由能最低(35.70 kcal·mol-1);群体的优势单倍型H1和H11突变体的自由能分别为-36.10和40.04 kcal·mol-1。gga-mir-1658基因不同单倍型成熟体种子区序列存在差异,在gga-mir-1658-5p存在"AUACCAU"、"AUACCAC"2种种子区序列,gga-mir-1658-3p共有"AACUCUG"、"AGCUGUG"、"AACUGUG"和"AGCUCUG"4种种子区序列。针对gga-mir-1658的预测靶基因的生物信息学分析显示,它们主要在基因表达调控、细胞凋亡、免疫系统的发育和B细胞激活等基本生物学过程中显著富集;此外,种子区变化可以影响gga-mir-1658成熟体对靶基因的选择。【结论】(1)gga-mir-1658基因种子区存在4个具有潜在生物学功能和表型效应的突变位点,可组成11种单倍型,其中H1(C C G–)和H11(G T G G)在北京油鸡、太行鸡和来航鸡为优势单倍型。(2)种子区突变影响gga-mir-1658基因前体二级结构的稳定性和靶基因的选择,可能是具有潜在表型效应的重要功能位点。
文摘The secondary structure of different Iβ cellulose was analyzed by a molecular dynamics sim- ulation with MARTINI coarse-grained force field, where each chain of the cellulose includes 40 D-glucoses units. Calculation gives a satisfied description about the secondary structure of the cellulose. As the chain number increasing, the cellulose becomes the form of a helix, with the diameter of screw growing and spiral rising. Interestingly, the celluloses with chain number N of 4, 6, 24 and 36 do show right-hand twisting. On the contrast, the celluloses with N of 8, 12, 16 chains are left-hand twisting. These simulations indicate that the cellulose with chain number larger than 36 will break down to two parts. Besides, the result indicates that 36-chains cellulose model is the most stable among all models. Furthermore, the Lennard-Jones potential determines the secondary structure. In addition, an equation was set up to analyze the twisting structure.
文摘One of the major obstacles facing the field of structural biology in the post genomic era is the inherent difficulty of analyzing the structure of membrane proteins under native conditions. The method of choice for studying such proteins is FTIR spectroscopy. Following the outbreaking of the severe acute respiratory syndrome （SARS） virus, in 2003, extensive work has been directed at elucidating the structure of the E transmembrane proteins of the SARS coronavirus. In this study, the secondary structure of the transmembrane a-helical bundles was analysised using the biophysical method site specific infrared dichroism （SSID）. Sixteen amino acids were isotopically labeled with （~3C=180） at different positions of the primary structure of the synthesized E protein CoV. The secondary structure was studied using Attenuated Total Internal Reflection （ATR） FTIR spectroscopy. Based on our findings, the presence of two possible H-bonding interactions between the carbonyl oxygen of two residues 26 and 31 （Phe and Leu） respectively with water molecules which may be trapped within the helix structure were postulatesed. These interactions may cause a change in this structure.
基金This work was supported by the National Natural Science Foundation of China （No.20373088）, the Program for Innovation Group （No.60321002）, the Innovative Project of Chinese Academy of Sciences （No.KJCX2-SW-w29）, and the National Key Project for Basic Research No.2006CB910302）. We thank Prof. Chih-chen Wang and Dr. Hui-min Ke in the Institute of Biophysics, Chinese Academy of Science, for the preparation of samples DsbC and G49R. We also thank Prof. Xiang-gang Qiu in the Institute of physics, Chinese Academy of Sciences, for help in FTIR measurement.
文摘The native protein structures in buffer solution are maintained by the electrostatic force as well as the hydrophobic force, salt ions play an important role in maintaining the protein native structures, and their effect on the protein stability has attracted tremendous interests. Infrared spectroscopy has been generally used in molecular structure analysis due to its fingerprint resolution for different species including macromolecules as proteins. However spectral intensities have received much less attention than the vibrational frequencies. Here we report that the spectral intensities of protein amide I band, the finger prints for the protein secondary structures, are very sensitive to the local electric field known as Onsager reaction field caused by salt ions. IR absorbance thermal titrations have been conducted for a series of samples including simple water soluble amino acids, water soluble monomeric protein cytochrome c and dimeric protein DsbC and its single-site mutant G49R. We found that at lower temperature range （10-20℃）, there exists a thermal activated salting-in process, where the IR intensity increases with a rise in the temperature, corresponding to the ions binding of the hydrophobic surface of protein. This process is absent for the amino acids. When further raising the temperature, the IR intensity decreases, this is interpreted as the thermal activated breaking of the ion-protein surface binding. Applying Van＇t Hoff plot to the thermal titration curves, the thermodynamic parameters such as AH and AS for salting-in and ion unbinding processes can be derived for various protein secondary structural components, revealing quantitatively the extent of hydrophobic interaction as well as the strength of the ion-protein binding.
