Baeyer-Villiger monooxygenases (BVMOs) are important biocatalysts to synthesize a series of valuable esters and lactones. Based on protein sequence alignment and crystal structure analysis, a nonconserved hinge which linked NADPH domain and FAD domain was speculated to play an important role in substrate recognition and catalytic oxidation process. Cyclohexanone monooxygenase (CHMO) was selected as a model. Mutants obtained by homologous replacement of the whole hinge almost completely lost its original catalytic activity, demonstrating that the overall hinge structure was of great importance. Some significant sites were identified to greatly affect the catalytic activity and stereoselectivity by alanine scanning mutagenesis, accompanied by enzyme activity assessments and chiral kinetic resolutions. Altering K153 decreased the activity of the enzyme but enhanced the stereoselectivity. Changing L143 site reduced stereoselectivity but had little effect on enzyme activity. Mutation at L144 site dramatically weakened both activity and stereoselectivity. Subsequently, these corresponding sites in phenylacetone monooxygenase were also illustrated to follow a similar rule, revealing a universal importance of these sites in the BVMO family. These results expanded our understanding of the structure-activity relationship of these enzymes and provided more proofs for future directed evolution of BVMOs.
Amino Acid Sequence ; Catalysis ; Mixed Function Oxygenases ; chemistry ; Models, Molecular ; NADP ; Oxidation-Reduction ; Oxygenases ; chemistry ; Protein Conformation ; Sequence Alignment ; Structure-Activity Relationship ; Substrate Specificity

Objective To investigate the risk factors,clinical prediction and intrapartum management of shoulder dystocia in non-macrosomia.Methods Totally 7 811 cases of vaginal delivery were retrospectively reviewed from Juanary 2009 to December 2013 in Shengjing Hospital.Shoulder dystocia was found in 11 cases (0.14％,11/7 811),including 1 case of macrosomia and l0 cases of non-macrosomia (shoulder dystocia group).Each non-macrosomia shoulder dystocia case was matched with 10 cases of normal delivery in the same week,which were selected randomly as the control group.The tendency and risk factors of shoulder dystocia in macrosomia and non-macrosomia were analyzed,and the following data between the two groups were compared,including the height of uterus fundus,abdominal circumference of the pregnant woman,the increasing of body mass index(BMI),fetal biparietal diameter (BPD),fetal femur length (FL),duration of every stage of labor,birth weight of the newborn,head circumference and chest circumference of the newborn,Apgar score.Results (1) There were 213 macrosomias among the 7 811 vaginal deliveries,with the incidence of 2.73％ (213/7 811).Only 1 shoulder dystocia was macrosomia (0.46％,1/213); while the other 10 cases were non-macrosomia (0.13％,10/7 598).(2) From 2009 to 2013,the macrosomia happened by 24 cases (2.32％,24/1 034),42 cases (3.61％,42/1 164),46 cases (2.60％,46/1 772),62 cases (3.01％,62/2 060),39 cases (2.19％,39/1781),respectively.The incidence of macrosomia had no significant difference among these 5 years (P＞0.05).The shoulder dystosia occurrence without macrosia in these 5 years were 1 case (0.10％,1/1 034),3 cases (0.26％,3/1 164),2 eases (0.11％,2/1 172),2 cases (0.10％,2/2 060),2 cases (0.11％,1/1 781),respectively.The incidence of shoulder dystocia without macrosomia had no significant difference among these 5 years (P＞0.05).(3) In the should dystocia group,5 cases were complicated with premature rupture of membrane (5/10),4 cases were mother≥ 35 years old (4/10),3 cases were multipara(3/10),3 cases had gestational diabetes mellitus(3/10),3 cases were occiput posterior during the first stage of labor(3/10),3 cases had prolonged second stage of labor (3/10) and 6 cases had routine lateral incision (6/10).In the control group,3 cases were complicated with premature rupture of membrane(3/10); 1 case was mother≥35 years old (1/10); 2 cases were multipara(2/10),3 cases had gestational diabetes mellitus (3/10),1 case had prolonged second stage (1/10) and 7 cases had routine lateral incision (7/10).(4) There were no significant difference in the height of uterus fundus,BMI,BPD,FL,and duration of the first stage of labor between the shoulder dystocia group and the control group (P＞0.05).Compared with the control group,the increasing of BMI [(6.8±3.1) vs (4.8± 1.4) kg/m2],the time of the second stage of labor[(86±65) vs (38±28) minutes] and abdominal circumference[(108±8) vs (101±7) cm] were significantly higher in the shoulder dystosia group (P＜0.05).(5)There were significant difference in the chest circumference of the newborn [(34.0±1.6) vs (32.2±1.9) cm] and the ratio of chest circumference to head circumference of the newborn [(0.99±0.03) vs (0.97±0.03)] between the two groups(P＜0.05).The 1-minute Apgar score of the newborn (7.4±2.8) was significantly lower than the control group (10.0±0.0) (P＜0.01).Clavicular fracture occurred in 3 newborns and brachial plexus injury occurred in 4 newborns in the shoulder dystosia group.Conclusion It is difficult to predict shoulder dystocia in non-macrosomia.Shoulder dystocia of non-macrosomia could be predicted by measurement of the head circumference,chest circumference,the ratio of chest circunfference to head circumference by using prenatal ultrasound.The risk factors may complicated with premature rupture of membrane,abnormal occiput position during the first stage of labor and prolonged second stage of labor.

Objective To investigate the relationship between blood pressure(BP) and prognosis in three different ischemic stroke subtypes.Methods The consecutive patients with a brain infarction proven on diffusion-weighted MRI who were hospitalized within 48 hours after stroke onset between April 2007 and April 2008 were registered.All subjects with acute ischemic stroke consecutively admitted to the neurological wards of the First Affiliated Hospital of Wenzhou Medical College,were registered in the Wenzhou Stroke Registry Program.Data were collected and coded at primary registration.The BP levels were studied during the initial 7 hospital days.Survival and dependency were assessed at 3 months.Outcomes were adjusted age,consciousness level,admission NIHSS score,the decline level of systolic BP,the decline level of diastolic BP,complication and so on. Logistic regression model was used to estimate the relationship between BP and prognosis.Results A U-shaped effect was observed in each subgroup between BP and prognosis.In the subgroups of atherothrombosis,cardioembolism and small artery disease,those who had a BP of 150/95 mm Hg(1 mm Hg=0.133 kPa)on admission,140/90 mm Hg on day 1-7 would have a better prognosis.In the subgroups of atherothrombosis and cardioembolism,the decrease of BP during the first 24 hours was the independent predictor of the death and disability at 3-month.In the atherothrombosis group,when the decrease of systolic BP during the first 24 hours was greater than 20 mm Hg,the risk of the death and disability at 3-month increased 4.44 times(OR 4.44,95%CI 1.70-11.59,P=0.002).In the atherothrombosis group,when the decrease of diastolic BP during the first 24 hours was greater than 10 mm Hg,the risk of the death/disability at 3-month increased 3.70 times(OR 3.70,95%CI 1.54-8.90.P=0.00).In the cardioembolism group,the risk increased respectively 7.98 times(OR 7.98,95%CI 1.34-47.66.P=0.026)and 6.68 times(OR 6.68.95%CI 1.55-28.79,P=0.01).In the subgroups of small artery disease,the decrease of BP during the first 24 hours was not the independent predictor of the death and disability at 3-month.Conclusions A U-shaped effect is observed in each subgroup between BP and prognosis.In the subgroups of atherothrombosis and cardioembolism,the decrease of BP during the first 24 hours is the independent predictor of the death and disability at 3-month.

[Objective] To learn about the genetic diversity,we studied the five X-chromosomal STR (X-STR) loci in Guangdong Han Nationality Groups.[Methods] The five Loci (DXS6803,DXS981,DXS6809,DXS6789,and DXS7132) were amplified in a pentaplex PCR reaction.PCR products were analyzed using capillary electrophoresis and ABI prism 3100 Genetic Analyzer,with GeneMapper ID 3.1 Analysis Software.[Results] A total of 363 individuals (181 unrelated male and 182 unrelated female) from Guangdong Han population were tested,54 alleles were observed for these loci.Polymorphism information content is 0.6935 ～ 0.8177.Power of discrimination in females was 0.8976 ～ 0.9562.Mean exclusion chance for X-STR in standard trios with daughters was 0.7805 ～ 0.8467.[Conclusion] The five loci in the multiplex system provide high polymorphism information for forensic identification and paternity testing,particularly for difficult paternity deficiency cases.

To improve the yield of insulin precursor (PI), we constructed a recombinant expression vector pPIC9K-PI and transformed it into Pichia pastoris GS115 using electroporation. After screening, a mutant strain CL012 with 12 copies was obtained on the YPDS plate containing 4.0 mg/mL G418. Then, the components of SNAREs (Soluble N-ethylmaleimide-sensitive factor attachment receptor proteins), SNC2 and SNC2-SSO2, were expressed in the strain CL012 to explore the effect of SNAREs on the yield of PI. In shake flask culture, the strains expressing SNC2 and SNC2-SSO2 yielded PI of 1.89 mg/L and 2.21 mg/L after methanol induction for 96 h, which were improved by 23.53% and 44.44% compared to that of strain CL012 (1.53 mg/L), respectively. Further, in a 5-L bioreactor, the yield of PI with strain CL012 was 53 mg/L for high-density fermentation, after 96 h of methanol induction, which was 34.64-fold higher than that of shake culture. The strains expressing SNC2 and SNC2-SSO2 yielded the PI of 64 mg/L and 78 mg/L, which were respectively increased by 20.75% and 47.17%, compared to that of strain CL012. This work indicated that SNAREs components promoted the secretion of PI to improve its heterologous expression in P. pastoris.

Neoadjuvant therapy has become the first choice for locally advanced esophageal carcinoma. Patients with post-neoadjuvant positive lymph node staging (ypN+) have poor prognosis, and there is no effective adjuvant therapy. Programmed death protein-1 (PD-1) antibody can obtain better clinical efficacy in the treatment of advanced esophageal cancer. The authors designed a multicenter, prospective, randomized controlled clinical trial of Toripalimab (PD-1 antibody) adjuvant therapy on esophageal squamous cell carcinoma patients with ypN+ after the treatment of neoadjuvant chemotherapy combined with surgical resection, in order to provide clinical practices for the adjuvant treatment of ypN+ patients.

Practice teaching and supervision has become an important part of hospital social work practice, and the handling and response to internship dilemma is the problem that every institution supervisor needs to think a-bout. Based on qualitative interview and text analysis, starting from the needs of intern, this paper found that students of social work major mainly faced the following dilemma in the process of hospital practice: role adapta-tion, the dilemma of professional practice, the dilemma of ethical relationship and practice burnout and so on. Combined with their own supervision experience, this paper put forward coping methods.

Objective:This study analyzes the expression level of miR-1180-3p and constructs the regulatory network of relevant ceRNA by integrating the DNA methylation and gene expression profile of hepatocellular carcinoma from the Cancer Genome Atlas (TCGA).Methods:Firstly, the expression level of miR-1180-3p in hepatocellular carcinoma and adjacent tissues was analyzed by TCGA database, and the differential expression of lncrna and mRNA was screened. Secondly, the LncBase database and the TargetScan database were used to predict the relationship between miR-1180-3p and lncRNA and mRNA, and the DNA methylation-mediated lncRNA was screened by the DNA methylation profile of lncRNA. Finally, Cytoscape software was used to construct miR-1180-3p relevant ceRNA network, and WebGestalt website was used to perform GO and KEGG analysis of related mRNA in ceRNA.Results:Compared with patients with low expression of miR-1180-3p (mean overall survival duration, 5.69 ± 0.35 years), patients with high expression of miR-1180-3p had shorter overall survival time (mean overall survival duration, 3.99 ± 0.47 years), indicating that the high expression of miR-1180-3p was hepatocellular carcinoma risk factor affecting the prognosis ( HR = 1.28, 95% CI = 1.1 ~ 1.5, P < 0.01). A miR-1180-3p related ceRNA regulatory network was constructed in this study, which contained 2 lncRNAs (F11-AS1 and LINC01511) and 37 mRNAs. Conclusion:This study has successfully constructed miR-1180-3p relevant ceRNA regulatory network, and DNA methylation-mediated F11-AS1 and F11-AS1/miR-1180-3p/C11of54 ceRNA regulatory axis has played an important role in the occurrence and development of hepatocellular carcinoma.

Microbial cell factories capable of producing valuable chemicals from renewable feedstocks provide a promising alternative towards sustainability. However, environmental stress remarkably affects the performance of microbial cell factories. By extending the chronological lifespan of microbial cells, the performance of microbial cell factories can be greatly improved. Firstly, an evaluation system for chronological lifespan and semi-chronological lifespan was established based on the changes in survival rates. Secondly, the addition of anti-aging drugs such as cysteine, carnosine, aminoguanidine and glucosamine increased the chronological lifespan of E. coli by 80%, 80%, 50% and 120%, respectively. Finally, we demonstrated that extending the chronological lifespan of E. coli increased the yield of metabolites produced by E. coli cell factories with endogenous (lactic acid and pyruvic acid) or exogenous (malic acid) metabolic pathway by 30.0%, 25.0%, and 27.0%, respectively. The strategy of extending chronological lifespan of E. coli provides a potential approach for enhancing the performance of microbial cell factories.
Escherichia coli/genetics* ; Lactic Acid ; Longevity ; Metabolic Engineering ; Metabolic Networks and Pathways

BACKGROUND: The relationship between quality of life at three months after lung cancer surgery and different surgical approaches is remains unclear. This study aimed to compare the quality of life of patients three months after uniportal and multiportal thoracoscopic lobectomy. METHODS: Data from patients who underwent lung surgery at the Department of Thoracic Surgery, Sichuan Cancer Hospital between April 2021 and October 2021 were collected. The European Organization for Research and Treatment of Quality of Life Questionnaire-Core 30 (EORTC QLQ-C30) and Quality of Life Questionnaire-Lung Cancer 29 (EORTC QLQ-LC29) were used to collect quality of life data of the patients. Potential confounding factors in the baseline data were included in a multivariate regression model for adjustment, and the quality of life of the two groups three months postoperatively was compared with traditional clinical outcomes. RESULTS: A total of 130 lung cancer patients were included, with 57 males (43.8%) and 73 females (56.2%), and an average age of (57.1±9.5) yr. In the baseline data of the two groups, there was a statistical difference in the number of chest drainage tubes placed (P<0.001). After adjustment with the regression model, at three months postoperatively, there were no significant differences in all symptoms and functional status scores between the two groups (all P>0.05). The multiportal group had longer surgery time (120.0 min vs 85.0 min, P=0.001), postoperative hospital stay (6.0 d vs 4.0 d, P=0.020), and a higher incidence of early ≥ grade 2 complications (39.0% vs 10.1%, P=0.011) compared to the uniportal group. CONCLUSIONS Patients undergoing uniportal and multiportal thoracoscopic lobectomy have similar quality of life at three months postoperatively. The uniportal group may have certain advantages in terms of traditional clinical outcome indicators such as operation time, postoperative hospital stay, and early postoperative complications.
Male ; Female ; Humans ; Lung Neoplasms/surgery* ; Quality of Life ; Thoracic Surgery, Video-Assisted/adverse effects* ; Pneumonectomy/adverse effects* ; Postoperative Complications/surgery* ; Retrospective Studies