The aim of this study was to prepare tandem multimeric proteins of BmSPI38, a silkworm protease inhibitor, with better structural homogeneity, higher activity and stronger antifungal ability by protein engineering. The tandem multimeric proteins of BmSPI38 were prepared by prokaryotic expression technology. The effects of tandem multimerization on the structural homogeneity, inhibitory activity and antifungal ability of BmSPI38 were explored by in-gel activity staining of protease inhibitor, protease inhibition assays and fungal growth inhibition experiments. Activity staining showed that the tandem expression based on the peptide flexible linker greatly improved the structural homogeneity of BmSPI38 protein. Protease inhibition experiments showed that the tandem trimerization and tetramerization based on the linker improved the inhibitory ability of BmSPI38 to microbial proteases. Conidial germination assays showed that His₆-SPI38L-tetramer had stronger inhibition on conidial germination of Beauveria bassiana than that of His₆-SPI38-monomer. Fungal growth inhibition assay showed that the inhibitory ability of BmSPI38 against Saccharomyces cerevisiae and Candida albicans could be enhanced by tandem multimerization. The present study successfully achieved the heterologous active expression of the silkworm protease inhibitor BmSPI38 in Escherichia coli, and confirmed that the structural homogeneity and antifungal ability of BmSPI38 could be enhanced by tandem multimerization. This study provides important theoretical basis and new strategies for cultivating antifungal transgenic silkworm. Moreover, it may promote the exogenous production of BmSPI38 and its application in the medical field.
Animals ; Antifungal Agents/pharmacology* ; Escherichia coli/metabolism* ; Proteins/metabolism* ; Protease Inhibitors/chemistry* ; Bombyx/chemistry* ; Saccharomyces cerevisiae/metabolism* ; Peptide Hydrolases

Proteases are widely found in organisms participating in the decomposition of proteins to maintain the organisms' normal life activities. Protease inhibitors regulate the activities of target proteases by binding to their active sites, thereby affecting protein metabolism. The key amino acid mutations in proteases and protease inhibitors can affect their physiological functions, stability, catalytic activity, and inhibition specificity. More active, stable, specific, environmentally friendly and cheap proteases and protease inhibitors might be obtained by excavating various natural mutants of proteases and protease inhibitors, analyzing their key active sites by using protein engineering methods. Here, we review the studies on proteases' key active sites and protease inhibitors to deepen the understanding of the active mechanism of proteases and their inhibitors.
Binding Sites ; Catalytic Domain ; Endopeptidases ; Peptide Hydrolases/genetics* ; Protease Inhibitors ; Proteins

Objective:To summarize and analyze the clinical data of Chinese patients with colony-stimulating factor 1 receptor (CSF1R)-related leukoencephalopathy, and clarify the phenotypic and genetic characteristics of Chinese patients.Methods:Medical history of patients with CSF1R-related leukoencephalopathy diagnosed from April 1, 2018 to January 31, 2021 in the department of neurology of 22 hospitals in China was collected, and scores of Mini-Mental State Examination (MMSE), Montreal Cognitive Assessment Scale (MoCA), magnetic resonance severity scale were evaluated. Group comparison was performed between male and female patients.Results:A total of 62 patients were included, and the male-female ratio was 1∶1.95. The age of onset was (40.35±8.42) years. Cognitive impairment (82.3%, 51/62) and motor symptoms (77.4%,48/62) were the most common symptoms. The MMSE and MoCA scores were 18.79±7.16 and 13.96±7.23, respectively, and the scores of two scales in male patients (22.06±5.31 and 18.08±5.60) were significantly higher than those in females (15.53±7.41 , t=2.954, P=0.006; 10.15±6.26, t=3.328 , P=0.003). The most common radiographic feature was bilateral asymmetric white matter changes (100.0%), and the magnetic resonance imaging severity scale score was 27.42±11.40, while the white matter lesion score of females (22.94±8.39) was significantly higher than that of males (17.62±8.74 , t=-2.221, P<0.05). A total of 36 CSF1R gene mutations were found in this study, among which c.2381T>C/p.I794T was the hotspot mutation that carried by 17.9% (10/56) of the probands. Conclusions:The core phenotypic characteristics of CSF1R-related leukoencephalopathy in China are progressive motor and cognitive impairment, with bilateral asymmetrical white matter changes. In addition, there exist gender differences clinically, with severer cognitive impairment and imaging changes in female patients. Thirty-six CSF1R gene mutations were found in this study, and c.2381T>C/p. I794T was the hotspot mutation.

To explore the mediating role of psychological resilience to childhood abuse and binge eating. This study assessed the childhood abuse, binge eating and psychological resilience of 3 453 middle school students in Harbin city, Heilongjiang Province. SPSS PROCESS macro program, combined with Bootstrap method, was employed to explore the mediating effect of psychological resilience. The incidence of middle school students experiencing at least one type of abuse in their childhood was 81.3% (2 807/3 453). Childhood abuse, psychological resilience and binge eating were all significantly different in terms of gender, household registration and whether they were only child (all P values<0.05). There was a significant positive correlation between childhood abuse and binge eating. Psychological resilience was negatively associated with childhood abuse and binge eating. Childhood abuse could not only directly predict the binge eating behavior of adolescents, but also could indirectly affect it via psychological resilience.

To explore the mediating role of psychological resilience to childhood abuse and binge eating. This study assessed the childhood abuse, binge eating and psychological resilience of 3 453 middle school students in Harbin city, Heilongjiang Province. SPSS PROCESS macro program, combined with Bootstrap method, was employed to explore the mediating effect of psychological resilience. The incidence of middle school students experiencing at least one type of abuse in their childhood was 81.3% (2 807/3 453). Childhood abuse, psychological resilience and binge eating were all significantly different in terms of gender, household registration and whether they were only child (all P values<0.05). There was a significant positive correlation between childhood abuse and binge eating. Psychological resilience was negatively associated with childhood abuse and binge eating. Childhood abuse could not only directly predict the binge eating behavior of adolescents, but also could indirectly affect it via psychological resilience.

Objective: To understand current situation and relations of childhood abuse and psychological capital, providing scientific basis for adolescent mental health promotion. Methods: Stratified cluster sampling method was used to select 1 894 students from junior high schools and senior high schools in Harbin. Questionnaire survey was conducted by using Childhood Trauma QuestionnaireShort Form (CTQ-SF) and Positive Psychological Capital (PPQ). Results: The total rate of childhood abuse among adolescents in the region was 98.3%. In addition to emotional abuse, scores of other dimensions of childhood abuse were higher for boys than for girls(P<0.01). Childhood abuse in rural area was higher than those in urban area except physical abuse(P<0.05). High psychological capital was observed among participants with boys higher in selfefficacy and resilience than that of girls(P<0.05). Psychological capital in urban students was higher than rural students(P<0.05). Except for the negative correlation between sexual abuse and resilience, all other dimensions of childhood abuse were negatively correlated with four dimensions of psychological capital. Stepwise regression analysis showed that emotional abuse, emotional neglect and physical neglect was negatively correlated with all dimensions of psychological capital(P<0.01); Sexual abuse showed negative association with selfefficacy and optimism(P<0.01). Conclusion Childhood abuse is closely related to psychological capital among adolescents in Harbin, suggesting exposure to childhood abuse might confer detrimental effects on psychological capital development.

Objective: To investigate the efficiency and safety of domestic tyrosine kinase inhibitor (TKI) dasatinib (Yinishu) as second-line treatment for patients with chronic myeloid leukemia in chronic phase (CML-CP). Methods: A retrospective analysis of clinical data of CML-CP patients who received domestic dasatinib as second-line treatment in the CML collaborative group hospitals of Hubei province from March 2016 to July 2018 was performed. The optimal response rate, the cumulative complete cytogenetic response (CCyR), the cumulative major molecular responses (MMR), progression free survival (PFS), event free survival (EFS) and adverse effects (AEs) of the patients were assessed at 3, 6 and 12 months of treatment. Results: A total of 83 CML-CP patients were enrolled in this study. The median follow-up time was 23 months. The optimal response rates at 3, 6 and 12 months in 83 CML-CP patients treated with dasatinib were 77.5% (54/71), 72.6% (61/75) and 60.7% (51/69), respectively. By the end of follow-up, the cumulative CCyR and MMR rates were 65.5% (55/80) and 57.1% (48/73), respectively. The median time to achieving CCyR and MMR was 3 months. During follow-up time, the PFS rate was 94.0% (79/83) and the EFS rate was 77.4% (65/83). The most common non-hematological AEs of dasatinib were edema (32.5%), rash itching (18.1%) and fatigue (13.3%). The common hematological AEs of dasatinib were thrombocytopenia (31.3%), leukopenia (19.3%) and anemia (6.0%). Conclusion Domestic dasatinib was effective and safe as the second-line treatment of CML-CP patients and it can be used as an option for CML-CP patients.

To explore the mediating role of psychological resilience to childhood abuse and binge eating. This study assessed the childhood abuse, binge eating and psychological resilience of 3 453 middle school students in Harbin city, Heilongjiang Province. SPSS PROCESS macro program, combined with Bootstrap method, was employed to explore the mediating effect of psychological resilience. The incidence of middle school students experiencing at least one type of abuse in their childhood was 81.3% (2 807/3 453). Childhood abuse, psychological resilience and binge eating were all significantly different in terms of gender, household registration and whether they were only child (all P values<0.05). There was a significant positive correlation between childhood abuse and binge eating. Psychological resilience was negatively associated with childhood abuse and binge eating. Childhood abuse could not only directly predict the binge eating behavior of adolescents, but also could indirectly affect it via psychological resilience.

Objective This study examined the prevalence of underweight and its related risk factors of community-dwelling patients with schizophrenia. Methods Five hundred and three community-dwelling patients with schizophrenia and 323 healthy controls were recruited in a cross-sectional study. Body mass index less than 18.5 was defined as underweight. Their demographic and clinical data including anthropometric data, plasma glucose and lipid parameters were collected. The Positive and Negative Syndrome Scale (PANSS) was used to assess patients' psychopathology. Results The prevalence of underweight was 9.9% (50/503) in schizophrenia patients versus 1.5% (5/323) in the control group ( P<0.01). Further logistic regression analysis showed that male ( OR=2.43, 95%CI:1.74~3.39), smoking behavior (OR=1.50, 95%CI: 1.21~1.86), hospitalization times (OR=1.18, 95% CI: 1.06~1.31), PANSS negative score (OR=1.09, 95% CI: 1.04~1.14) were significant predictors for underweight (all P<0.05). Conclusion The prevalence of underweight is higher in Chinese patients with schizophrenia than in the general population. Some demographic and clinical variables are risk factors for underweight in schizophrenia.

Objective To compare cognitive function and alexithymia between the schizophrenic patients treated with chlorpromazine and those treated with clozapine. Methods The patients with schizophrenia in stable condition that received maintenance treatment either with chlorpromazine or clozapine and normal control subjects were recruited (n=24 per group). Neuropsychological tests, including Digit Vigilance Test, Controlled Oral Word Association Test, Trail Making Test-A&B (TMT-A&B), Animal Naming Test, Stroop Color-Word Test, Block Design and Spatial Span Test were used to assess the participant’s cognitive function. The twenty-item Toronto Alexithymia Scale (TAS-20) were used to evaluate the participant’s alexithymia. Results The significant differences were found between the patients and the controls in all items of cognitive function and all factor scores of TAS-20 (P<0.05). There were no significant differences in all items of cognitive function and all factor scores of TAS-20 between chlorpromazine group and clozapine group (P>0.05). Multivari? ate linear regression analysis showed that in the clozapine group, attention function associated with the total score of TAS (β=-0.20, P<0.05), executive function associated with TAS factor 1 (β=-0.26, P=0.03), spatial function associated with TAS factor 2 (β=-0.24, P<0.01). In the chlorpromazine group, attention function associated with TAS factor 2 (β=-1.24, P<0.01), executive function associated with TAS factor 2 (β=-0.33, P=0.02). Conclusions Patients with schizophrenia in maintenance period have widely cognitive impairment and alexithymia, both of which are related to each other.