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Arginine kinase (AK) is a key enzyme in energy metabolism of invertebrates and plays an important regulatory role in the life activities such as growth and development, nutrition utilization, immune resistance and stress response. Arginine kinase of Bombyx mori (BmAK) is related to the energy balance and anti-NPV process, but there is little research on its molecular structure and enzymatic properties. We cloned the ORF sequence of BmAK gene, and analyzed chromosomal localization, genomic structure, mRNA structure, secondary and tertiary structure. Phylogenetic analysis indicated that AK was highly conserved in evolution. Soluble recombinant BmAK was obtained by prokaryotic expression, and purified by Ni-NTA affinity chromatography. The circular dichroism spectroscopy showed that BmAK contained α-helix structures, and its α-helix structures were relatively stable in the pH range between 5 and 10. Enzyme activity analysis showed that the optimum temperature of BmAK was 30 ℃ and the optimum pH of BmAK was 7.5. The optimal temperature of BmAK was 25 ℃. Between 15 ℃ and 30 ℃, the structure and activity of BmAK was relatively stable. The structure of BmAK was relatively stable at pH 7.0. Our findings reveal the structure and function of BmAK to develop novel green safe and environmentally friendly insecticides.
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Protein tyrosine phosphatase (PTP, EC 3.1.3.48) specifically catalyzes the removal of phosphate groups from phosphorylated tyrosine residues, resulting in protein dephosphorylation, thus regulates life activities such as cell growth, proliferation, differentiation and immunization. Protein tyrosine phosphatase h of Bombyx mori (BmPTP-h) is involved in the replication of nucleopolyhedrovirus (NPV) in Bombyx mori, but the structure and properties of BmPTP-h are little known so far. In this study, the coding sequence of BmPTP-h gene was cloned from the midgut of Bombyx mori, and its genomic structure, mRNA structure, sequence signature, secondary structure and the state in solution were analyzed. Homologous amino acid sequences alignment analysis indicated that BmPTP-h had a high similarity to PTP sequences of numbers of insect NPVs, implying that they may have a common ancestor and similar function. We constructed a prokaryotic expression vector, expressed and obtained the soluble recombinant BmPTP-h in Escherichia coli at 25 ℃, and purified BmPTP-h using Ni-NTA affinity chromatography. Gel filtration analysis showed that BmPTP-h was able to form aggregate and monomer in solution. Circular dichroism spectroscopy analysis showed that the recombinant BmPTP-h contained α-helix structure. Increasing temperature resulted in the unfolding of the α-helix structure of BmPTP-h and the decrease of the α-helix structure content of BmPTP-h. These studies provide a basis to better study the structure and regulation mechanism of BmPTP-h.
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Objective To explore the mediating effects of college students＇ online social support on the relationship between online self-disclosure and internet altruistic behavior.Methods Using cluster random sampling,the self-disclosure questionnaire for Chinese undergraduates with their internet friend,the online social support scale of adolescence and the internet altruistic behavior scale of undergraduates were applied to 351 college students.Results The score of college students＇ online self-disclosure,online social support and internet altruistic behavior were respectively(2.22±0.45),(2.93±0.59) and (2.04±0.46).The college students＇ online self-disclosure,online social support and internet altruistic behavior were positively correlated with each other (P＜0.01).College students＇ online self-disclosure not only predicted internet altruistic behavior directly,but also predicted internet altruistic behavior indirectly through online social support.The direct effects and indirect effects were 0.200 and 0.265.The ratio of the mediating effect on the total effect was 57％.The fit index of mediation model was x2/df=2.87,RMSEA =0.07,CFI =0.94,TLI =0.93,GFI=0.92,IFI=0.94,NFI=0.91.Conclusion The college students＇ online self-disclosure not only affects the interuet altruistic behavior directly,but also can take the same effect on online social support indirectly.
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Basic helix loop helix (bHLH) transcription factor plays an important role in biological processes. Bmsage is a class of bHLH transcription factor highly expressed in the silk gland of Bombyx mori, which is not only involved in the developmental regulation of the silk gland cells at the embryonic period, but also plays a crucial regulatory role during the synthesis of silk protein. However, currently, much of the property and structure of Bmsage is still remained unknown. To study the property, structure and biological role of Bmsage, we constructed several prokaryotic expression vectors of Bmsage fused with NusA, MBP, SUMO, Trx and His tags, respectively, then screened and determined the best soluble expression vector and condition of Bmsage in Escherichia coli combining with the induction temperature and IPTG concentration, and further purified the recombinant Bmsage by Ni-column affinity chromatography according to the established expression condition and characterized its secondary structure using circular dichroism spectra. The results showed that NusA and MBP could significantly enhance the soluble expression of Bmsage in E. coli, but it was difficult to separate Bmsage from these tags. SUMO could not only increase the soluble expression of Bmsage in E. coli to a certain degree, but also be effectively separated from Bmsage. Other tags did not effectively promote the soluble expression of Bmsage in E. coli. Circular dichroism spectra showed that the purified Bmsage had well-defined α-helix structure in solution, indicating that SUMO may promote the correct folding of Bmsage into native-like structure. These work not only establish a foundation for further study of the property, structure and function of Bmsage, but also provide a reference for the expression and purification of other similar proteins.
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Objective To examine the association between the polymorphisms of brain-derived neurotrophic factor (BDNF)gene with heroin dependence.Methods Genomic DNA was isolated from the venous blood leukocytes of 308 unrelated patients with heroin dependence and 31 7 healthy individuals.Seven single nucleotide polymorphisms (SNPs)were genotyped using MassARRAY system.Data were analyzed using HaploView 4.0 and SPSS 20.0 software.Results There was a significant difference in the genotype frequency of rs6265 between heroin dependence group and healthy control group (χ2 =1 5.1 5 1,P =0.001).The rs6265 G allele was significantly higher than in controls (χ2 =9.864,P =0.002,OR =1.429,95% CI =1.143 -1.786).Furthermore,there was also a significant difference in the genotype frequency of rs13306221 between heroin dependence group and control group (χ2 =7.699,P =0.006).The rs13306221 G allele was significantly higher than in controls (χ2 =7.137,P =0.008,OR =0.539,95% CI =0.340-0.853).Strong linkage disequilibrium (LD)was observed in one block (D’> 0.9;r 2 >0.8),and significantly less G-G haplotype frequency of block 1 (χ2 =4.546;P =0.033)was found in heroin dependence group. Conclusion Our findings support the role of BDNF rs6265 and rs13306221 polymorphisms in heroin dependence and may guide future studies to identify other genetic risk factors for heroin dependence.
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Objective To study the effect of dopamine receptors on neurological and physiological activities. Methods Dopamine D1 receptor gene (DRD1) knockout mice and dopamine D3 receptor (DRD3) gene knockout mice were introduced, and double gene knockout mice were bred in our lab.Seven SPF male mice in each group were used in this experiment.The food intake, water intake, body weight gain for 24 hours were tested on the age of 30 d, 50 d, and 70 d and were compared with those of wild type mice.Results DRD1 gene and DRD3 gene showed significant effect on the body weight in mice in age of 21 day and 35 day, but at the age of 90 day, the differences became insignificant among the mice of various genetypes.Conclusions Dopamine may effect on the foraging and satiety in newborn mice through regulating the hypothalamic-pituitary-adrenal ( HPA ) axis activity, and finally leads to a reduced body weight gain in newborn mice and puppies during lactation.Furthermore, DRD1 gene and DRD3 gene may influence on body weight of newborn mice through regulating mothers’ lactation, lead to a lower body weight at ablactation, and compensatory increase of body weight after ablactation.Our results provide a substantial foundation for studying the function and interaction of DRD1 and DRD3 genes.
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Objective To test whether adolescent rats are more sensitive than adult rats to morphine' s rewarding actions and locomotor activating effects. Methods Conditioned place preference(CPP) procedure was used to examine drug-induced reward and locomotor activity. Adolescent ( Postnatal day 35 ) and adult ( P67 ) male rats received either saline or morphine (3 mg/kg) for 6 days and the locomotor response in the drug paired chamber was recorded during conditioned training. CPP effects were tested 24h after the last training. Results In adolescent rats, morphine CPP effect was not present [ saline group (20. 89 ±31. 14) s, morphine group (90. 75 ± 27.91)s, P = 0. 15], however, morphine CPP effect occurred in adult rats[saline group (31.5 ± 41. 24) s, morphine group(266. 13 ± 32. 26) s, P < 0. 001 ]. Adult rats presented higher CPP score than adolescent rats(P < 0.001). The locomotor response was significantly higher in day 3 ～ 6 than day 1 in the adolescent treated with morphine (P<0.05 or <0. 01). The locomotor response was significantly higher in day 2-6 than day 1 in the adolescent treated with morphine(P<0. 05 or < 0. 01 ) .which suggested that both adolescent and adult rats exhibited similar behavioral sensitization to morphine administration. Two-way ANOVA indicated that the main effect of age was not significant(F_(1,14) =0. 33, P = 0. 57) ,the interaction effect of age x treat was not significant(F_(5,70) = 0.85, P = 0.52) .there was no age difference between adolescent and adult rats in behavioral sensitization effects. Conclusion Compared with adult rats, adolescent rats don' t present morphine CPP, but show the similar morphine behavioral sensitization.
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Objective: To test whether depressive college students had sociotropic self-schemata. Methods: A total of 500 college students were surveyed by CES-D and SAS, and 77 college students were chosen for further experiment: 24 with depressive mood, 12 with anxious mood, 17 with both depressive and anxious mood, and 24 normal ones. Sociotropic adjectives were strictly developed, and the 4 groups (depressive vs. anxious vs. depressive and anxious vs. normal) were given Self-referent Encoding Task. Results: ①The depressive group showed significantly less positive adjectives and more negative adjectives for self-descriptive than the normal group (P
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Objective To investigate the genetic polymorphism of DXS8378 STR locus of chromosome X in Chinese Lisu,Pumi and De'ang populations in Yunnan and construct relative standard allelic ladders.Methods After being amplified by PCR,different STR allelic fragments were isolated from the PAG electrophoresis.The STR allelic fragments were extracted by kit and reamplified by PCR to obtain purified allelic fragments.Next,the purified allelic fragments were subcloned individually into the PUC plasmid vectors,and the size and structure of the inserts were confirmed by the analysis of their DNA sequences.Then we transfected it into competent E.coli DH5?TM cells,and finally,the recombinant plasmids DNA with the inserts were used as template for reamplification to generate the standard ladders.Results The standard allelic ladder for DXS8378 locus was obtained,with which the genetic polymorphisms of DXS8378 locus in three Chinese populations in Yunnan were studied.Conclusion The standard ladder made by this method is excellent,and DXS8378 is powerful for forensic practice in Chinese population.