The main commercial production of fructooligosaccharides (FOS) comes from enzymatic transformation using sucrose as substrate by microbial enzyme fructosyltransferase. A fructosyltransferase genomic DNA was isolated from Aspergillus niger QU10 by PCR. The nucleotide sequence showed a 1 941 bp size, and has been submitted to GenBank (KF699529). The cDNA of the fructosyltransferase, containing an open reading frame of 1 887 bp, was further cloned by RT-PCR. The fructosyltransferase gene from Aspergillus niger was functionally expressed both in Escherichia coli and Pichia pastoris GS 115. The highest activity value for the construction with the α-factor signal peptide reached 431 U/mL after 3 days of incubation. The recombinant enzyme is extensively glycosylated, and the active form is probably represented by a homodimer with an apparent molecular mass of 200 kDa as judged from mobility in seminative PAGE gels. The extracellular recombinant enzyme converted sucrose mostly to FOS, mainly 1-kestose and nystose, liberating glucose. FOS reached a maximal value and represented about 58% of total sugars present in the reaction mixture after 4 h reaction. The results suggest that the availability of recombinant Pichia pastoris as a new source of a FOS-producing enzyme might result of biotechnology interest for industrial application.
Aspergillus niger ; enzymology ; genetics ; Base Sequence ; Cloning, Molecular ; DNA, Complementary ; Escherichia coli ; Fungal Proteins ; genetics ; metabolism ; Glycosylation ; Hexosyltransferases ; genetics ; metabolism ; Molecular Sequence Data ; Molecular Weight ; Pichia ; Sucrose ; metabolism ; Trisaccharides ; metabolism

Objective To understand the short-term effect of practice skill training for general practitioners in Guangdong province and to provide scientific basis for improving the training strategy. Methods According to the training outline (Outline of job-transfer training for general practitioner in primary health institutions (trial))of the Ministry of Health,job-transfer training was provided for a total of 451 primary care doctors in economically undeveloped cities for one year. All trainees were investigated before and after training by questionnaire,which were designed by ourselves and included practical skill level and satisfaction degree.SPSS 13.0 software was used to do statistical analysis and data were processed by description analysis,t test,ANOVA and correlation analysis. Results Average self-evaluation scores of practical skills were 136.37 ±21.74 and 169.39 ±17.12 before and after training with significant differences (t=29.028,P=0.000). Scores of all types of practical skills after training were significantly higher than those before training (P<0.000 1). Total satisfaction rate was 91.6%and students were not well satis-fied with training environment and training guarantee. Fac-tors influencing the effect of training included educational background,professional title,doctors' de-sire of training,whether taking part in the general practitioner training or not. The effect of practical training was positively correlated with total satisfaction (r=0.162,P=0.037). Conclusions Remark-able results of practice skill training have been achieved. In the future,more chances for practice should be provided for trainees and contents of skill training should be designed according to the need of community health service,and more attention should be paid to the training project management.

ObjectiveTo summarize the method and experience of the model of rat heterotopic heart transplantation and to improve the rate of success of the experiment.Methods120 heterotopic heart transplantations were performed by Ono's way,including 60 prepared transplantations and 60 formal transplantations.ResultsRate of success of prepared transplantations was 45％,and formal transplantation was 85％.Mean time of formal operation was (74.3±17.1)min,removal time of the donor heart was (7.9±4.2)min,ischemia time was (29.6±10.3) min and the survival time of allograft was (11.1±5.7)days.5 of the recipients had a long-term survival,and the maximum weight was 740 g.ConclusionsGood surgical technique and rigorous perioperative management is very important in setting up the rat model of heterotopic heart transplantation.

OBJECTIVETo observe the clinical efficacy and safety on moderate and severe persistent allergic rhinitis treated with acupuncture.

METHODSSixty-six patients of moderate and severe persistent allergic rhinitis were randomized into an acupuncture group (34 cases) and a western medication group (32 cases). In the acupuncture, group, acupuncture was applied to Dazhui (GV 14), Feishu (BL 13), Pishu (BL 20), Ganshu (BL 18) and Shenshu (BL 23) in the prone, retained for 20 min; then in the supine, at Baihui (GV 20), Yintang (GV 29), yingxiang (LI20) Taichong (LR 3) and Hegu (LI 4), retained for 20 min. Acupuncture was given once every two days, three times a week, continuously for 8 weeks. In the western medication group, cetirizine hydrochloride was taken orally, 10 mg each time, once every day, continuously for 8 weeks. Separately, before treatment, after the treatment of 1 and 2 months and in 1 month after treatment, the total nasal symptom score (TNSS), the scores in the emotion rating scale for Ganzangxiang of TCM (ERSG) and the rhinoconjunctivitis quality of life questionnaire (RQLQ) were observed in the patients of the two groups. The clinical efficacy was compared between the two groups.

RESULTS(1) For TNSS, the results after 1 and 2 months treatment and in 1 month after treatment were all, reduced as compare with that before treatment separately in the two groups (P < 0.05, P < 0.01) The result after 2 months treatment was lower than that after 1 month treatment in the acupuncture group (P < 0.05). In 1 month after treatment, the result in the acupuncture group was lower than that in the western medication group (P < 0.05). (2) For ERSG, the score after 2 months treatment was lower than that before treatment in the two groups (both P < 0.05). The score after 2 months treatment in the acupuncture group was lower than that in the western medication group (P < 0.05). (3) For RQLQ, the score after 1 month treatment was lower than that before treatment and the score after 2 months treatment was lower than that after 1 month treatment in the two groups (all P < 0.05). The score after 1 and 2 months treatment and in 1 month after treatment in the acupuncture group was lower than that in the western medication group separately (all P < 0.05). (4) The total effective rate was 91.2% (31/34) in the acupuncture group and was 90.6% (29/32) in the western medication group, without significant difference between the two groups (P > 0.05).

CONCLUSIONAcupuncture is the safe and effective intervention on moderate and severe persistent allergic rhinitis. Compared with the western medicine group, the efficacy in the acupuncture group presents much more advantageous at its durability.

Acupuncture Points ; Acupuncture Therapy ; Adult ; Female ; Humans ; Male ; Middle Aged ; Quality of Life ; Rhinitis, Allergic ; diagnosis ; therapy ; Young Adult

Objective The purpose of this study was to investigate the expression of human mitochondrial transcription termi-nation factor-3 ( hMTERF3) in non-small cell lung cancer ( NSCLS) and to analyze its clinicopathological significance. Methods The paraffin block samples used in this study included 65 cases of NSCLC and 32 cases of normal alveolar epithelial tissues. We determined the expressions of hMTERF3 in NSCLC and normal alveolar epithelial tis-sues by immunohistochemistry, calculate the survival rate using the Kaplan-Meier method, and analyzed the risk factors affecting the prognosis of NSCLC using the Cox Proportional Hazard Model. Results In the 65 cases of NSCLC, 31 ( 47. 69%) showed positive expression of hMTERF3. The total survival time was significantly shor-ter in the patients with a high than in those with a low hMTERF3 ex-pression ([30.39±3.35] vs [57.61±7.12] mo, P<0.05). The riskfactors affecting the prognosis of NSCLC included positive expression of hMTERF3 (HR=3.302, 95% CI:1.598-6.905) and lymph node metastasis (HR=4.052, 95% CI: 1.212-12.398). Conclusion hMTERF3 is overexpressed in NSCLC. Highly expressed hMTERF3 and lymph node metastasis reduce the survival time of NSCLC patients, suggesting that hMTERF3 may be a potential bio-marker for the prognosis of NSCLC.

Objective To analyze quantitatively the histological components, and to observe the morphological changes of the transition zone in benign hyperplastic prostates (BPH) and adult normal prostates. Methods The quantitative morphometry of 40 BPH specimens obtained from suprapubic transvesical prostatectomy and 10 normal prostatic specimens (controls) were performed with computer-assisted image analysis system,and all the samples were stained with hematoxylin and eosin (HE) and the Masson trichrome.The area percentages of stromal (smooth muscle and connective tissue),glandular epithelial and luminal components,the average areas of acini,lumens and epithelia,and the heights of epithelial cells were quantitated,respectively. Results ①The mean area percentages of different components between control group and BPH group were as follows: smooth muscles,(23.83?8.53)% vs (35.35?8.33)% ( P 0.05);epithelia,(26.26? 7.45)% vs (17.76?4.61)% (P0.05).②The average areas of acini, epithelia and lumens of control group and BPH group were ( 0.087?0.028)mm 2 vs (0.062?0.030)mm 2 (P

Objective To investigate the association between hepatitis B virus (HBV)genotype, the mutations in HBV basic core gene promoter(BCP), pre C/C gene region and treatment response to interferon (IFN)α-1b. Methods Hepatitis B e antigen (HBeAg)-positive chronic hepatitis B (CHB) patients were treated with IFNα-Ib for 6 months and were followed up for 6 months after the end of treatment. Restriction Fragment Length Polymorphism (RFLP) was used for determining HBV genotype. HBV DNA was amplified by polymerase chain reaction (PCR) and analyzed for BCP and pre C/C gene region by sequencing. Measurement data were compared using t test and analysis of variance. Enumeration data were compared using chi-square test, Fisher exact probability test.Logistic regression analysis was utilized for multi-factor analysis. Results There were 39 patients who completed the treatment and follow up in this study. At the end of treatment, 16(41.0%) patients showed response to the IFNα-lb treatment. At the end of follow-up, four out of 16 patients who achieved on treatment response relapsed. Among 3a patients, 29 (74.4 %) were infected with genotype B and 10 (25. 6%) with genotype C. The treatment response rates were not significant different between the groups with different genotypes. The double mutation pattern (T1762/A1764) was found in eight (20. 5%) patients. The response rates to IFNα-lb treatment were not significant different between the group with and without double mutation pattern. A1896 mutation was detected in eight patients at baseline. Three of them became HBeAg negative at the end of treatment and returned to HBeAg positive during follow-up. The non-lyphocyte epitope mutations, L60V and I97L, were found in 15 patients (38. 5%) and 14 patients (35.9%), respectively. At the end of follow-up, the patients with 60V had a significantly lower HBeAg seroconversion rate and HBV DNA undetectable rate compared to the patients with 60L (Fisher exact probability test; P = 0.0126 and 0.0069,respectively). The HBV DNA undetectable rates in the patients with 97I were significantly lower than those in patients with 97L both at the end of treatment and the end of follow-up (Fisher exact probability test; P= 0.0484 and 0. 0024, respectively). Logistic regression analysis results showed that there was no association between the above viral mutations and the treatment response to IFNαlb. Conclusions There is no association between HBV genotype, BCP double mutation pattern and IFN-α treatment response. The non-lyphocyte epitope mutations, L60V and I97L, may have impact on IFN-α treatment response.

In this study, we investigated the mechanism of transformation by Bacillus subtilis strain 168 by proteomic analysis. B. subtilis strain 168 was able to stereoselectively transform cis-propenylphosphonic acid (cPPA) to fosfomycin. The maximal fosfomycin production was 816.6 microg/mL after two days cultivation, with a conversion rate of 36.05%. We separated the whole cellular proteins by two-dimensional gel electrophoresis (2-DE) method, and 562 protein spots were detected in the presence of cPPA in the medium, while 527 protein spots were detected in the absence of cPPA. Of them, 98 differentially expressed protein spots were found. Among them, 52 proteins were up-regulated whereas 20 were down-regulated in the presence of cPPA in the medium, and 26 induced at the presence of cPPA. The differentially expressed proteins were analyzed by combined MS and MS/MS methods. Eighty protein spots, including 45 up-regulated proteins, 17 down-regulated proteins, and 18 induced by cPPA were identified. Based on the results of proteomic analysis, we postulated two steps of transformation: in the first step, cPPA was hydrated to 2-hydroxypropylphosphonic acid; in the second step, 2-hydroxypropylphosphonic acid was transformed to fosfomycin via a dehydrogenation reaction.
Bacillus subtilis ; genetics ; growth & development ; metabolism ; Bacterial Proteins ; metabolism ; Biotransformation ; Fosfomycin ; metabolism ; Organophosphorus Compounds ; metabolism ; Proteome ; metabolism ; Proteomics

Objective: To investigate the efficacy of sequential therapy with telbivudine in the treatment of HBeAg-positive chronic hepatitis B (CHB) patients with partial response after a standard course of interferon therapy. Methods: A retrospective cohort study was performed for 58 HBeAg-positive CHB patients with partial response at the end of interferon therapy (48-60 weeks) from January 2009 to December 2013. According to whether telbivudine was used sequentially or withdrawn at the end of the course of treatment, the patients were divided into telbivudine sequential therapy group and withdrawal group, and the two groups were compared with in terms of biochemical, virological, and serological response rates. The chi-square test, the t-test, and the non-parametric test were used based on data type. Results: A total of 58 patients were enrolled in this study, with 31 in the telbivudine sequential therapy group and 27 in the withdrawal group. At 12 and 24 weeks after interferon therapy ended, the telbivudine sequential therapy group had a significantly higher HBeAg clearance rate than the withdrawal group (22.6%/29.0% vs 0%/3.7%, P < 0.05). At week 48 of follow-up, the telbivudine sequential therapy group had a significantly higher combined response rate than the withdrawal group (22.6% vs 0%, P = 0.015). Among the 31 patients in the telbivudine sequential therapy group, 11 had an increase in creatine kinase during the administration of telbivudine. No patient in either group experienced serious adverse reactions during follow-up, such as muscular soreness, myositis, peripheral neuropathy, renal dysfunction, and liver function decompensation. Conclusion In HBeAg-positive CHB patients with partial response to interferon therapy, sequential therapy with telbivudine can increase serological HBeAg clearance rate and combined response rate at week 48, and it is safe in HBeAg-positive CHB patients achieving partial response at the end of interferon therapy.