Chain-length dependence of the kinetics of the hyaluronan hydrolysis catalyzed by bovine testicular hyaluronidase

Hyaluronan (HA) has various biological functions that are strongly dependent on its chain length. In some cases, as in inflammation and angiogenesis, long and short chain-size HA effects are antagonistic. HA hydrolysis catalyzed by hyaluronidase (HAase) is believed to be involved in the control of the balance between longer and shorter HA chains. Our studies of native HA hydrolysis catalyzed by bovine testicular HAase have suggested that the kinetic parameters depend on the chain size. We thus used HA fragments with a molar mass ranging from 8x10(2) g mol(-1) to 2.5x10(5) g mol(-1) and native HA to study the influence of the chain length of HA on the kinetics of its HAase-catalyzed hydrolysis. The initial hydrolysis rate strongly varied with HA chain length. According to the Km and Vm/Km values, the ability of HA chains to form an efficient enzyme-substrate complex is maximum for HA molar masses ranging from 3x10(3) to 2x10(4) g mol(-1). Shorter HA chains seem to be too short to form a stable complex and longer HA chains encounter difficulties in forming a complex, probably because of steric hindrance. The hydrolysis Vm values strongly suggest that as the chain length decreases the HAase increasingly catalyses transglycosylation rather than hydrolysis. Finally, two HA chain populations, corresponding to HA chain molar masses lower and higher than approximately 2x10(4) g mol(-1), are identified and related to the bi-exponential character of the model we have previously proposed to fit the experimental points of the kinetic curves.     

Cd2+、Cu2+胁迫对黑藻(Hydrilla verticillata)的生长及光合荧光特性的影响

以黑藻(Hydrilla verticillata)为供试材料,采用Cd2+和Cu2+等两种重金属分别在5个浓度梯度水平下的河砂水培方法,研究Cd2+或Cu2+不同浓度胁迫对黑藻株高、生物量、成活率和叶绿素含量的影响,以及对黑藻叶片最小荧光(Fo)、最大荧光(Fm),PSⅡ最大量子产率(QYmax,)、稳态下的PSⅡ反应中心关闭程度(1-Qp_Lss)、稳态下的非光化学淬灭(NPQ_Lss)等光合荧光参数及其荧光成像的影响,探讨各个参数分别随镉、铜浓度递增的变化规律。研究结果表明,Cd2+胁迫下黑藻的株高显著下降,说明Cd2+可能对黑藻叶片的维管束结构产生伤害作用;Cd2+和Cu2+胁迫对黑藻鲜重均无显著影响,说明与水生植物自由水含量存在一定关系;Cd2+和Cu2+胁迫均使黑藻干重显著下降,其中Cu2+胁迫对黑藻干重的影响更显著,说明Cu2+胁迫对黑藻累积生物量的影响远大于Cd2+;Cd2+和Cu2+胁迫下叶绿素各值均呈下降趋势,而Cu2+胁迫对叶绿素的影响更大,说明Cu2+对黑藻叶绿体的毒害比Cd2+更大。随着Cd2+或Cu2+胁迫浓度梯度的增加,黑藻的叶绿素荧光参数(Fo、Fm、QYmax)呈显著下降趋势,(1-Qp_Lss)呈上升趋势,而NPQ_Lss先上升后下降。黑藻在不同重金属胁迫下的生理指标、荧光参数及成像特征等方面所表现出的变化差异性,反映出同等浓度下黑藻受重金属胁迫的影响程度为:Cd2+胁迫Cu2+胁迫;黑藻可以在Cu2+浓度低于1 mg/L的环境下具有正常的光合活性,可推测将黑藻用于低浓度Cu污染水域的修复;在Cu2+浓度达3 mg/L以上环境下黑藻即无法长时间生存,可推测黑藻可以作为Cd污染水环境的指示种。     

Modulation of hematology changes by polymorphism of glutathione S-transferase M1 and T1

Workers in the petroleum distribution trades experience relatively low-level exposures to gasoline vapors whose consequences have not been fully elucidated. The purpose of this study was to investigate changes in the hematological parameters among filling station workers who were occupationally exposed to gasoline. The target group for the study consisted of 41 workers from eight filling stations of Shiraz (south of Iran). The control group consisted of 27 healthy subjects matched for age and sex from general population. The complete blood count analysis was done in one laboratory. Using PCR-based method, the genotypes of glutathione S-transferase T1 (GSTT1) and M1 (GSTM1) were determined. Workers were divided into three exposure groups according to employment history: duration less than 1 year, 1-5 years, and more than 5 years. Comparison was performed using Kruskal-Wallis test. In the individuals with the presence of both GSTT1 and GSTM1 functional alleles, comparison between four exposure groups revealed no significant difference for studied hematological variables. There were statistically significant differences between study groups, with only one functional allele, either GSTT1 or GSTM1, for relative number of lymphocytes (chi(2)=9.147, df=3, P=0.027) and neutrophils (chi(2)=9.951, df=3, and P=0.019), and absolute number of lymphocytes (chi(2)=9.135, df=3, and P=0.028), and RBC (chi(2)=10.586, df=3, and P=0.014). These findings could indicate the possible protective effect of concurrent presence of GSTM1 and GSTT1 enzymes on the hematopoietic system of filling station workers.     

Modeling Neisseria meningitidis B metabolism at different specific growth rates

Neisseria meningitidis is a human pathogen that can infect diverse sites within the human host. The major diseases caused by N. meningitidis are responsible for death and disability, especially in young infants. At the Netherlands Vaccine Institute (NVI) a vaccine against serogroup B organisms is currently being developed. This study describes the influence of the growth rate of N. meningitidis on its macro-molecular composition and its metabolic activity and was determined in chemostat cultures. In the applied range of growth rates, no significant changes in RNA content and protein content with growth rate were observed in N. meningitidis. The DNA content in N. meningitidis was somewhat higher at the highest applied growth rate. The phospholipid and lipopolysaccharide content in N. meningitidis changed with growth rate but no specific trends were observed. The cellular fatty acid composition and the amino acid composition did not change significantly with growth rate. Additionally, it was found that the PorA content in outer membrane vesicles was significantly lower at the highest growth rate. The metabolic fluxes at various growth rates were calculated using flux balance analysis. Errors in fluxes were calculated using Monte Carlo Simulation and the reliability of the calculated flux distribution could be indicated, which has not been reported for this type of analysis. The yield of biomass on substrate (Y(x/s)) and the maintenance coefficient (m(s)) were determined as 0.44 (+/-0.04) g g(-1) and 0.04 (+/-0.02) g g(-1) h(-1), respectively. The growth associated energy requirement (Y(x/ATP)) and the non-growth associated ATP requirement for maintenance (m(ATP)) were estimated as 0.13 (+/-0.04) mol mol(-1) and 0.43 (+/-0.14) mol mol(-1) h(-1), respectively. It was found that the split ratio between the Entner-Doudoroff and the pentose phosphate pathway, the sole glucose utilizing pathways in N. meningitidis, had a minor effect on ATP formation rate but a major effect on the fluxes going through for instance the citric-acid cycle. For this reason, we presented flux ranges for underdetermined parts of metabolic network rather than presenting single flux values, which is more commonly done in literature.     

Combined analysis of group recorded feed intake and individually recorded body weight and litter size in mink

In the mink industry, feed costs are the largest variable expense and breeding for feed efficient animals is warranted. Implementation of selection for feed efficiency must consider the relationships between feed efficiency and the current selection traits BW and litter size. Often, feed intake (FI) is recorded on a cage with a male and a female and there is sexual dimorphism that needs to be accounted for. Study aims were to (1) model group recorded FI accounting for sexual dimorphism, (2) derive genetic residual feed intake (RFI) as a measure of feed efficiency, (3) examine the relationship between feed efficiency and BW in males (BWM) and females (BWF) and litter size at day 21 after whelping (LS21) in Danish brown mink and (4) investigate direct and correlated response to selection on each trait of interest. Feed intake records from 9574 cages, BW records on 16 782 males and 16 875 females and LS21 records on 6446 yearling females were used for analysis. Genetic parameters for FI, BWM, BWF and LS21 were obtained using a multivariate animal model, yielding sex-specific additive genetic variances for FI and BW to account for sexual dimorphism. The analysis was performed in a Bayesian setting using Gibbs sampling, and genetic RFI was obtained from the conditional distribution of FI given BW using genetic regression coefficients. Responses to single trait selection were defined as the posterior distribution of genetic superiority of the top 10% of animals after conditioning on the genetic trends. The heritabilities ranged from 0.13 for RFI in females and LS21 to 0.59 for BWF. Genetic correlations between BW in both sexes and LS21 and FI in both sexes were unfavorable, and single trait selection on BW in either sex showed increased FI in both sexes and reduced litter size. Due to the definition of RFI and high genetic correlation between BWM and BWF, selection on RFI did not significantly alter BW. In addition, selection on RFI in either sex did not affect LS21. Genetic correlation between sexes for FI and BW was high but significantly lower than unity. The high correlations across sex allowed for selection on standardized averages of animals’ breeding values (BVs) for RFI, FI and BW, which yielded selection responses approximately equal to the responses obtained using the sex-specific BVs. The results illustrate the possibility of selecting against RFI in mink with no negative effects on BW and litter size.     

Immobilization of lipase from Candida rugosa on Eupergit C supports by covalent attachment

The present study compares the results of three different covalent immobilization methods employed for immobilization of lipase from Candida rugosa on Eupergit^® C supports with respect to enzyme loadings, activities and coupling yields. It seems that method yielding the highest activity retention of 43.3% is based on coupling lipase via its carbohydrate moiety previously modified by periodate oxidation. Study of thermal deactivation kinetics at three temperatures (37, 50 and 75 °C) revealed that the immobilization method also produces an appreciable stabilization of the biocatalyst, changing its thermal deactivation profile. By comparison of the t_1/2 values obtained at 75 °C, it can be concluded that the lipase immobilized via carbohydrate moiety was almost 2-fold more stable than conventionally immobilized one and 18-fold than free lipase. The immobilization procedure developed is quite simple, and easily reproduced, and provides a promising solution for application of lipase in aqueous and microaqueous reaction system.     

Improved side-chain prediction accuracy using an ab initio potential energy function and a very large rotamer library

Accurate prediction of the placement and comformations of protein side chains given only the backbone trace has a wide range of uses in protein design, structure prediction, and functional analysis. Prediction has most often relied on discrete rotamer libraries so that rapid fitness of side-chain rotamers can be assessed against some scoring function. Scoring functions are generally based on experimental parameters from small-molecule studies or empirical parameters based on determined protein structures. Here, we describe the NCN algorithm for predicting the placement of side chains. A predominantly first-principles approach was taken to develop the potential energy function incorporating van der Waals and electrostatics based on the OPLS parameters, and a hydrogen bonding term. The only empirical knowledge used is the frequency of rotameric states from the PDB. The rotamer library includes nearly 50,000 rotamers, and is the most extensive discrete library used to date. Although the computational time tends to be longer than most other algorithms, the overall accuracy exceeds all algorithms in the literature when placing rotamers on an accurate backbone trace. Considering only the most buried residues, 80% of the total residues tested, the placement accuracy reaches 92% for chi(1), and 83% for chi(1 + 2), and an overall RMS deviation of 1 A. Additionally, we show that if information is available to restrict chi(1) to one rotamer well, then this algorithm can generate structures with an average RMS deviation of 1.0 A for all heavy side-chains atoms and a corresponding overall chi(1 + 2) accuracy of 85.0%.     

Hierarchical clustering of 54 races and strains of the mulberry silkworm,Bombyx mori L: Significance of biochemical parameters

Summary A detailed analysis was undertaken to test the efficacy of hierarchical agglomerative clustering (UPGMA method) in grouping the races and strains of the mulberry silkworm, Bombyx moti L., and to ascertain the importance of biochemical parameters in the clustering process. The analysis was based on data from two rearing seasons with 54 selected races/strains of different geographic origin and varying yield potentials. The results indicate that seven clusters can be realised with yield parameters alone, whereas the inclusion of biochemical parameters in clustering resulted into two broad groups: one having all the breeds with high cocoon weight and shell weight, the other having all the low-yielding silkworm strains both from India and from other countries. Further sub-grouping under these two groups highlights genetical differences associated with the differentiation of various groups of races in temperate and tropical areas as well as their significance for silkworm breeding. Estimates of all ten variables were further subjected to quick clustering and the results showed that cluster 5, constituted by 38 lowyielding strains of India, China and Europe, had the highest values of the final cluster centre for amylase and the effective rate of rearing (ERR), while clusters 1 and 4 had the highest values for invertase and alkaline phosphatase. The evolutionary aspect of the genetic channelisation of silkworm races from various countries is discussed against the background of differences in the biochemical parameters and yield variables.     

不同油松种源光合和荧光参数对水分胁迫的响应特征

采用盆栽控水试验方法,设置正常水分(T1,田间持水量的70％-80％)、轻度胁迫(T2,田间持水量的50％-60％)和严重胁迫(T3,田间持水量的30％-40％)3个水分梯度,对6个油松(Pinus tabulaeformis Carr.)种源(陕西洛南LN,陕西桥山 QO,山西灵空山LK,辽宁千山QN,河北雾灵山WL和山西芦芽山LY)的光合参数和叶绿素荧光参数进行比较,探讨种源间光合和水分利用特征的差异及其与气孔导度和荧光参数的关系.结果表明,3个水分梯度下6个种源的净光合速率(Pn)、蒸腾速率(Tr)、气孔导度(Gs)、水分利用效率(WUE)、原初光能转换效率(Fv/Fm)和PSⅡ潜在活性(Fv/Fo)差异均极显著(P＜0.01),在T1和T3下,各种源的实际光量子效率φPSⅡ和电子传递速率ETR差异显著(P＜0.05).在T2下,LK和WL的Pn达到最大,具有轻度胁迫下提高光合生产力的倾向.在T3下,QN种源的Pn和WUE较高(除LK的WUE最高外),而WL的Pn和WUE较低(除LK的Pn最低外);6个种源中QN的Pn、φPSⅡ和ETR最大,LY和QO的光合生产力仅次于QN,而LN、LK和WL的Pn、Gs、φPSⅡ和ETR比较低,其光合生产力及光量子产量受干旱胁迫影响较大.LK的WUE在各个水分梯度下均显著高于其它种源(P＜0.05).在T3下,各种源通过降低Tr提高WUE.在T1下,气孔限制是影响油松Pn的主要因素;在T2和T3下,Pn和WUE均与气孔导度、PSⅡ反应中心的光量子产量和电子传递速率密切相关.     

颅内压参数联合血清caveolin-1、AQP-4对高血压脑出血患者术后预后不良的预测价值

摘要 目的：探讨颅内压参数联合血清小窝蛋白-1（caveolin-1）、水通道蛋白4（AQP-4）对高血压脑出血（HICH）患者术后预后不良的预测价值。方法：选择2020年1月至2022年1月河北省胸科医院收治的106例HICH患者,术后随访3个月,根据格拉斯哥预后（GOS）评分将患者分为预后良好组（55例）,预后不良组（51例）。术后监测颅内压参数[压力反应指数（PRx）、平均颅内压波幅（MWA）、20 mmHg阈值下颅内压剂量（Dicp20）],检测血清caveolin-1、AQP-4水平。多因素Logistic回归分析HICH患者术后预后不良的因素。受试者工作特征曲线（ROC）分析颅内压参数联合血清caveolin-1、AQP-4预测HICH患者术后预后不良的价值。结果：预后不良组PRx、MWA、Dicp20以及血清caveolin-1、AQP-4水平高于预后良好组（P＜0.05）。低术前格拉斯哥昏迷评分（GCS）评分、高PRx、高Dicp20、高caveolin-1、高AQP-4是HICH患者术后预后不良的危险因素（P＜0.05）。联合PRx、Dicp20、caveolin-1和AQP-4预测HICH患者术后3个月预后不良的的曲线下面积为0.823,大于PRx、Dicp20、caveolin-1和AQP-4单独预测。结论：高PRx、Dicp20、caveolin-1、AQP4是HICH患者术后预后不良的危险因素,联合颅内压参数PRx、Dicp20及血清caveolin-1、AQP4预测HICH患者术后预后不良具有较高的价值。