[Methylthioadenosine phosphorylase and p16 as surrogate diagnostic markers for CDKN2A homozygous deletion in brain tumors].

Objective: To examine whether immunohistochemistry of methylthioadenosine phosphorylase (MTAP) and p16 could be used to predict the CDKN2A status in various brain tumors. Methods: A total of 118 cases of IDH-mutant astrocytomas, 16 IDH-wildtype glioblastoma, 17 polymorphic xanthoastrocytoma (PXA) and 20 meningiomas diagnosed at Xuanwu Hospital, Capital Medical University, Beijing, China from November 2017 to October 2023 were collected and analyzed. The CDKN2A status was detected by using fluorescence in situ hybridization or next-generation sequencing. Expression of MTAP and p16 proteins was detected with immunohistochemistry. The association of loss of MTAP/p16 expression with CDKN2A homozygous/heterozygous deletion was examined. Results: Among the 118 cases of IDH-mutant astrocytoma, 13 cases showed homozygous deletion of CDKN2A. All of them had no expression of MTAP while 9 cases had no expression of p16. Among the 16 cases of IDH wild-type glioblastoma, 6 cases showed homozygous deletion of CDKN2A. All 6 cases had no expression of MTAP, while 3 of these cases had no expression of p16 expression. Among the 17 PXA cases, 4 cases showed homozygous deletion of CDKN2A, and the expression of MTAP and p16 was also absent in these 4 cases. Among the 20 cases of meningiomas, 4 cases showed homozygous deletion of CDKN2A. Their expression of MTAP and p16 was also absent. Among the four types of brain tumors, MTAP was significantly correlated with CDKN2A homozygous deletion (P<0.05), with a sensitivity of 100%. However, it was only significantly correlated with the loss of heterozygosity (LOH) of CDKN2A in astrocytomas (P<0.001). P16 was associated with CDKN2A homozygous deletion in IDH-mutant astrocytoma and PXA (P<0.001), but not with the LOH of CDKN2A. Its sensitivity and specificity were lower than that of MTAP. Conclusions: MTAP could serve as a predictive surrogate for CDKN2A homozygous deletion in adult IDH-mutant astrocytoma, PXA, adult IDH-wildtype glioblastoma and meningioma. However, p16 could only be used in the first two tumor types, and its specificity and sensitivity are lower than that of MTAP.

[Clinicopathological and molecular genetic characteristics of adult IDH wild-type diffuse gliomas].

Objective: To analyze the clinicopathological and molecular features and prognostic implications of adult isocitrate dehydrogenase wild type (IDH-wt) diffuse gliomas. Methods: A total of 87 cases of adult IDH-wt diffuse gliomas from 2016 to 2020 in Xuanwu Hospital of Capital Medical University were retrospectively collected. The clinicopathological characteristics and prognosis were analyzed. Molecular characteristics were also analyzed using Sanger sequencing and next generation sequencing. Results: There were 53 males and 34 females, aged from 19 to 78 years (mean 53 years). Histopathologically, there were 63 (72.4%) glioblastomas, 16 (18.4%) anaplastic astrocytomas, six (6.9%) diffuse astrocytomas, and one (1.1%) each of anaplastic oligodendrocytoma, and anaplastic oligodendroglioma. Common molecular genetic changes in IDH-wt gliomas included TERT promoter mutation which was found in 60 cases (69.0%); MGMT promoter methylation in 43 cases (49.4%); EGFR mutation in 38 cases (43.7%); PTEN mutation in 35 cases (40.2%) and TP53 mutation in 32 cases (36.8%). In addition, PDGFRA mutation was detected in 17 cases (19.5%), CDK4 amplification in 15 cases (17.2%) and MDM2 amplification in 11 cases (12.6%). In IDH-wt diffuse gliomas, there was no significant difference in the overall survival between TERT promoter, EGFR, PTEN, TP53, PDGFRA, CDK4, MDM2 mutations and the wild-type, since these gene mutations could co-occur in any case (P>0.05). Also there was no significant difference in the overall survival between the WHO grade Ⅱ/Ⅲ gliomas and glioblastoma patients with these gene mutations (P>0.05). Conclusions: TERT promoter, EGFR, PTEN, TP53, PDGFRA, CDK4 and MDM2 gene mutations are common molecular genetic changes in adult IDH-wt gliomas, and are associated with poor prognosis. It is suggested that these genes are potentially useful for predicting the prognosis and should be tested in adult IDH-wt gliomas.

[High-grade gliomas with H3 G34R mutation: a clinicopathological study].

Objective: To analyze the clinicopathological features and probable mechanisms of high-grade gliomas with H3 G34R mutation. Methods: Five cases of high-grade gliomas with H3 G34R mutation were collected at Xuanwu Hospital, Capital Medical University, Beijing, China, from 2016 to 2019. The clinical and pathological data for each case was retrospectively reviewed. Results: The 5 patients (2 males and 3 females) aged from 15 to 45 years (mean 23 years), and had a history of headache or motor weakness. Four of them were younger than 20 years of age. Magnetic resonance imaging showed that the lesions of 3 cases were seen separately in frontal lobe, parietal lobe or temporal lobe, 1 case involved both frontal lobe and parietal lobe, and otherwise multiple lobes were involved in 1 case. Contrast enhancement could be observed in 2 cases. Pathological examination showed that glioblastoma was the most common entity, with or without primitive neuronal component. All 5 cases showed that H3 G34R was diffusely positive in tumor nuclei with ATRX loss. Moreover, p53 was overexpressed in 4 cases. None of them showed Olig2 expression. Two patients showed disease progression after surgery at 18 months and 24 months, respectively. The latter of the two deceased 3 months after tumor progression. Conclusions: The clinicopathological and molecular genetics features of high-grade gliomas with H3 G34R mutation have relatively similar clinicopathological and genetic features, and more commonly seen in young adults (vs. older adults). Thus, these tumors may be discussed further as a distinct tumor entity.

Peginterferon is preferable to entecavir for prevention of unfavourable events in patients with HBeAg-positive chronic hepatitis B: A five-year observational cohort study.

At present, the long-term effects of pegylated interferon-α (PEG-IFN-α) and entecavir (ETV) are controversial. Studies directly compared the long-term outcomes of these two drugs have not been completed. This study was designed to compare the clinical outcomes of PEG-IFN-α vs ETV therapy in Chinese patients with chronic HBV infection. From September 2008 to December 2016, a large, observational, open-label, prospective cohort study of HBeAg-positive patients with CHB who received PEG-IFN-α or ETV therapy was carried out at the Second Affiliated Hospital of Chongqing Medical University. Cumulative incidences of unfavourable events were calculated with respect to treatment type. Based on the REACH-B model, we compared the observed incidence of hepatocellular carcinoma (HCC) with the expected incidence in each group. PEG-IFN-α-treated patients showed a lower cumulative incidences of unfavourable events and cirrhosis than those treated with ETV (P = .031; P = .044, respectively). Impact factor exploration indicated that treatment type and platelet count are significantly associated with the occurrence of unfavourable events. Based on the REACH-B model, a lower observed cumulative incidence of HCC was observed in PEG-IFN-α-treated patients than predicted (P = .038). However, there was no significant difference of the cumulative HCC incidence between the observed and the predicted cases for ETV-experienced patients (P = .36). Treatment with PEG-INF-α leads to a lower incidence of unfavourable events including cirrhosis and HCC than ETV in patients with HBV. Treatment type and baseline platelet count may be two important factors associated with the long-term clinical outcomes of patients with CHB.

Fibre type-dependent response of broiler muscles to dietary antioxidant supplementation for oxidative stability enhancement.

The influence of dietary antioxidants and quality of oil on the oxidative and physico-chemical properties of chicken broiler breast and thigh meat stored was studied in either an oxygen-enriched (HiOx: 80% O2/20% CO2) or an air-permeable polyvinylchloride (PVC) packaging system during retail display at 2-4°C for up to 14 and 7 d, respectively. Broilers were fed on a diet with either a low-oxidised (peroxide value (POV) 23 meq O2/kg) or a high-oxidised (POV 121 meq O2/kg) oil, supplemented with or without an algae/selenium-based antioxidant with organic minerals, for 42 d. Lipid and protein oxidation, myofibrillar protein profile and purge loss were analysed. In both packaging systems, lipid oxidation (thiobarbituric acid-reactive substances [TBARS]) was inhibited by up to 65% and 57% in chicken breast and thigh, respectively, with an antioxidant-supplemented diet compared to those without. In both breast and thigh samples, protein sulfhydryls and water-holding capacity (purge loss) were better protected by the antioxidant dietary treatment, regardless of oil quality. Thigh muscles had up to sevenfold greater TBARS formation and more myosin heavy chain losses compared to breast samples. Antioxidant supplementation was more protective against lipid oxidation and water-holding capacity in the group fed on high-oxidised oil compared to those fed on low-oxidised oil. The results suggest that dietary antioxidants can minimise the negative impact of oxidised oil on broiler meat quality, and this protection was more pronounced for thigh than breast muscle, indicating inherent variations between muscle fibre types.

Augmentation of water-holding and textural properties of breast meat from oxidatively stressed broilers by dietary antioxidant regimens.

1. The impact of dietary antioxidants and degree of oil oxidation on textural attributes of chicken broiler breast meat stored in oxygen-enriched, air-permeable polyvinylchloride and skin packaging systems during retail display at 2-4°C for up to 21 d was assessed. 2. Broilers were fed on diets either with a low-oxidised oil (peroxide 23 mEq O2/kg) or with a high-oxidised oil (peroxide 121 mEq O2/kg), with or without an algae-based antioxidant and organic mineral antioxidant supplement for 42 d. 3. Fatty acids and radical scavenging activities of the diets were estimated. Meat colour, pH, myofibrillar protein profile and textural traits were measured. 4. Diets with high-oxidised oil reduced stearic, linoleic and linolenic acid content compared to low-oxidised oil samples, regardless of antioxidant supplementation. Meat colour and pH varied among dietary treatments throughout storage. Meat samples from the antioxidant dietary group, irrespective of oil oxidation level, had lower amounts of purge and cooking losses compared to the unsupplemented diets. For all packaging systems, meat shear force was significantly higher for broilers fed on high-oxidised diets. 5. The results demonstrate that dietary antioxidant supplementation can minimise the negative impact of oxidised oil on the quality of broiler meat packaged in different atmospheric environments.

[Research on cholera vaccine injection in the later period of the Republic of China].

During the period of the Republic of China, cholera was one of the government-mandated infectious diseases. In response to the cholera epidemic, the Beijing municipal government and the Health Bureau established the Beijing special epidemic prevention committee to guide and supervise the implementation of epidemic prevention measures. The Beijing municipal government and the Health Bureau cooperated with medical institutions to implement cholera vaccine injection at their locations, stations, gates, schools and other public places, as well as personal residences. They strictly defined the vaccinated population and vaccination methods, and adopted the vaccination mode of combining voluntary injection and compulsory injection, effectively controlled the spreading of the cholera. The public benefit, solidarity and flexibility of its epidemic prevention work are of certain reference significance to the epidemic prevention work today and even in the future.

[Effect of peer education on knowledge, attitude and practices towards schistosomiasis among primary school students in endemic foci of Wuhan City].

OBJECTIVE: To evaluate the effect of peer education on knowledge, attitude and practices towards schistosomiasis control among primary school students in endemic foci of Wuhan City. METHODS: Yucai Hankou Primary School in Jiang' an District, Wuhan City was selected in 2021, and all students at grades 4 and 5 were sampled using a cluster sampling method to receive peer education about schistosomiasis. The changes of knowledge, attitude and practices towards schistosomiasis control were compared before and after peer education to evaluate the effect of peer education on knowledge, attitude and practices towards schistosomiasis control. RESULTS: The overall awareness of schistosomiasis control knowledge and the overall proportion of correct attitudes towards schistosomiasis control and correct schistosomiasis-related behaviors increased from 51.50%, 93.70% and 92.99% before peer education to 86.50%, 98.98% and 98.72% after peer education among primary school students, respectively (χ2=149.457, 21.692 and 20.691, all P values < 0.05). The overall awareness of schistosomiasis control knowledge and the overall proportion of correct attitudes towards schistosomiasis control and correct schistosomiasis-related behaviors were 49.19%, 92.20% and 92.72% among Grade 4 primary school students and 53.83%, 95.21% and 93.28% among Grade 5 primary school students prior to peer education (χ2 = 1.214, 2.034 and 0.096, all P values > 0.05), and increased to 75.93%, 98.09% and 97.59% among Grade 4 primary school students and 97.16%, 99.87% and 99.87% among Grade 5 primary school students after peer education, respectively (χ2 = 40.798, 9.572, 7.207, 133.194, 9.678 and 14.926, all P values < 0.05). The overall awareness of schistosomiasis control knowledge and the overall proportion of correct attitudes towards schistosomiasis control and correct schistosomiasis-related behaviors were 51.25%, 76.92% and 77.97% among male primary school students and 51.80%, 94.42% and 95.70% among female primary school students prior to peer education, the differences in the overall proportion of correct attitudes towards schistosomiasis control and correct schistosomiasis-related behaviors between male primary school students and female primary school students were significant (χ2 = 30.462 and 33.416, both P values < 0.05), and increased to 86.23%, 98.25% and 97.79% among male primary school students and 86.83%, 99.85% and 99.85% among female primary school students after peer education (χ2 = 0.081, 3.529 and 3.335, all P values > 0.05), respectively. CONCLUSIONS: Peer education is effective to improve the knowledge, attitude and practices towards schistosomiasis control among primary school students in endemic foci of Wuhan City, which may be more effective to improve the awareness of schistosomiasis control knowledge and proportion of correct schistosomiasis-related behaviors among primary school students at high grades.

Complement inhibition enables renal allograft accommodation and long-term engraftment in presensitized nonhuman primates.

Protection against humoral injury mediated by donor-specific antibodies (DSA), also known as accommodation, may allow for long-term allograft survival in presensitized recipients. In the present study, we determined the role of complement in renal allograft accommodation in donor skin-presensitized nonhuman primates under conventional immunosuppression. Donor skin allografts were transplanted to presensitized recipients 14 days prior to renal transplantation. Renal allografts not receiving any immunosuppressive treatment developed accelerated rejection with predominantly humoral injury, which was not prevented using conventional cyclosporine (CsA) triple therapy. Inhibition of complement activation with the Yunnan-cobra venom factor (Y-CVF) successfully prevented accelerated antibody-mediated rejection and resulted in successful accommodation and long-term renal allograft survival in most presensitized recipients. Accommodation in this model was associated with the prevention of the early antibody responses induced against donor antigens by complement inhibition. Some antiapoptotic proteins and complement regulatory proteins, including Bcl-2, CD59, CD46 and clusterin, were upregulated in the surviving renal allografts. These results suggest that the complement inhibition-based strategy may be valuable alternative in future clinical cross-match positive or ABO-incompatible transplantation.

A novel fiber optic sensor to monitor beef meat emulsion stability using visible light scattering.

Accurate control of the meat emulsification process for a consistent product quality entails the development of an on-line optical sensor technology to determine the optimum chopping end-point yielding minimum cooking loss and a fine texture. Previous studies suggested that light backscatter measurements can be used to monitor physical-chemical changes during emulsification in comminuted meat products if appropriate spacing between the emitting and detecting optical fibers is used. Light backscatter intensity from beef emulsions manufactured with different fat/lean ratio (0.075, 0.250, and 0.330) and chopping duration (2, 5, and 8min) were obtained using a dedicated fiber optic prototype. Optical measurements were collected at three radial distances (2, 2.5, and 3mm) from the light source using a fiber optic spectrometer (300-1100nm). Light backscatter intensity decreased logarithmically with increasing fiber optic spacing. Light propagation through the emulsion decreased significantly with increasing chopping duration and fat concentration. Cooking loss increased with increasing fat/lean ratio and with under- or over-chopping. The maximum emulsion stability was observed at 5min of chopping. Several optically derived parameters were found to be significantly correlated with fat loss during cooking. Typically, those correlations were observed to increase with decreasing fiber distance. Based on these findings, an optical configuration is proposed that would compensate for the emulsion heterogeneity, maximizing the existing correlation between the optical signal and the emulsion quality metrics.

Changes in the physiochemical, microbial, and sensory characteristics of fresh pork sausage containing rosemary and green tea extracts during retail display.

The effects of adding combinations of rosemary (R: 1500, 2000, 2500 ppm) and green tea (G: 100, 200, 300 ppm) extracts in combination with synthetic antioxidants on the physiochemical, microbial, and sensory characteristics of fresh pork sausage were evaluated. R and G improved (P < .05) oxidative stability as evidenced by lower TBARS. R2500 and G300 had fewer PPC than the control at d 7, 14, and 21 of storage. Consumer acceptability scores were greater (P < .05) in sausages with R and G when compared to the control, and the majority of the R and G treatments were liked by 98% of the respondents. Treatment combinations of at least R2000 and G200 were described by positive drivers of liking such as spice-complex, ginger, nutmeg, rosemary flavors and aromas and lower scores for descriptors such as rancid, fruity, and off-flavor/odor. This research demonstrates that rosemary and green tea extracts improved the keeping quality of fresh pork sausage under simulated retail display.

Frozen storage stability of antioxidant-treated raw restructured beef steaks made from mature cows.

Previous research has shown that beef quality decreased with the age of cattle. In this study, beef trimmings from nine mature cows (n=9), equally representing three animal age groups (2-4, 6-8, and 10-12yr), were restructured into steaks formulated with propyl gallate, alone or in combination with a beefy flavoring agent, to enhance palatability and stability during 6months of frozen storage at -29°C. Lipid oxidation, rancidity, and loss of beefy flavor in restructured steaks during extended storage were reduced by propyl gallate. The beefy flavoring agent inclusion masked mature, forage-fed beef off-flavors, intensified beefy flavor, and improved steak tenderness, juiciness and cooking yield. Thus, the combination of propyl gallate and beefy flavoring offers an effective means to enhance the palatability and storage stability of restructured beef prepared from mature cows.

Prolonged cardiac allograft survival in presensitized rats after a high activity Yunnan-cobra venom factor therapy.

UNLABELLED: Complement-dependent antibody-mediated acute humoral rejection is the major obstacle of clinical transplantation across ABO incompatibility and human leukocyte antigen presensitization. We previously demonstrated that Yunnan-cobra venom factor (Y-CVF) could almost completely abrogate complement activity and successfully prevent hyperacute rejection in some xenotransplant models without any obvious toxicity. In this study we investigated whether depletion of complement by Y-CVF prevented acute humoral allograft rejection in presensitized rats thereby prolonging graft survival. METHODS: Presensitization was achieved in Lewis rats by sequential grafting of three full-thickness skin pieces from Brown Norway rats. Serum cytotoxic alloantibody titers were determined by a modified in vitro complement-dependent microcytotoxicity assay. After presensitization, each Lewis rat received a heterotopic Brown Norway cardiac allograft. Fifteen recipients were divided into two groups: (1) no treatment control (n = 7); (2) Y-CVF therapy group (86 u/kg, IV, day -1) (n = 8). After cessation of the heart beat, allograft rejection was confirmed by pathologic as well as IgG and C3 immunohistochemical examinations. RESULTS: The mean graft survival time was significantly prolonged to 99.50 +/- 38.72 hours among rats that received Y-CVF vs 12.71 +/- 13.94 hours in nontreated controls (P < .001). Upon pathological and immunohistochemical examination, acute humoral rejection was mainly exhibited in the control group, whereas acute cellular rejection was mainly displayed in the Y-CVF therapy group. CONCLUSIONS: Our study demonstrated that complement depletion by Y-CVF significantly inhibited acute humoral allograft rejection in presensitized rats. As a therapeutic immunointervention tool for complement, Y-CVF has shown potential efficacy across ABO incompatible and positive cross-match barriers.

Effect of dietary ractopamine on tenderness and postmortem protein degradation of pork muscle.

Twenty-four finishing pigs with a mean starting weight of 82kg were assigned to two dietary regimens: (1) a corn-soybean meal basal diet (control; n=12), and (2) the basal diet supplemented with 20ppm ractopamine HCl (RAC; n=12). After 28-30 days on the feeding trial, pigs were slaughtered, and the growth and carcass characteristics were measured. Furthermore, the 3rd-13th rib section of longissimus muscle was excised at 48h postmortem, sliced into 19-mm thick chops, vacuum packaged, stored at 2°C, and subjected to Warner-Bratzler shear force (WBSF) and electrophoretic tests after 2, 4, 7, 10, 14, and 21 days (postmortem). RAC feeding increased (P<0.01) pig carcass weight and percent lean, but it also increased the day-2 muscle WBSF by 20% (P<0.01). The shear force difference between control and RAC pig muscles gradually decreased and vanished by day 10 (P>0.05) when both muscle groups became more tender. The muscle from RAC-fed pigs exhibited a slower protein degradation rate than muscle from the control animals, notably for proteins in the 15-45kDa range. The results suggested that the tenderness difference between ractopamine-treated and control pig muscles was related to the proteolysis rate, and could be diminished with adequate postmortem ageing.

In vitro activities of small peptides from snake venom against clinical isolates of drug-resistant Mycobacterium tuberculosis.

The recent re-emergence of tuberculosis, especially the multidrug-resistant cases, has highlighted the importance of screening effective novel drugs against Mycobacterium tuberculosis. In this study, the in vitro activities of small peptides isolated from snake venom were investigated against multidrug-resistant M. tuberculosis. Minimum inhibitory concentrations (MICs) were determined by the Bactec TB-460 radiometric method. A small peptide with the amino acid sequence ECYRKSDIVTCEPWQKFCYREVTFFPNHPVYLSGCASECTETNSKWCCTTDKCNRARGG (designated as vgf-1) from Naja atra (isolated from Yunnan province of China) venom had in vitro activity against clinically isolated multidrug-resistant strains of M. tuberculosis. The MIC was 8.5 mg/l. The antimycobacterial domain of this 60aa peptide is under investigation.

An activator of blood coagulation factor X from the venom of Bungarus fasciatus.

A specific activator of blood coagulation factor X was purified from the venom of Bungarus fasciatus by gel filtration and by ion-exchange chromatography on a Mono-Q column (FPLC). It consisted of a single polypeptide chain, with a mol. wt of 70,000 in reducing and non-reducing conditions. The enzyme had an amidolytic activity towards the chromogenic substrates S-2266 and S-2302 but it did not hydrolyse S-2238, S2251 or S-2222, which are specific substrates for thrombin, plasmin and factor Xa, respectively. The enzyme activated factor X in vitro and the effect was Ca2+ dependent with a Hill coefficient of 7.9. As with physiological activators, the venom activator cleaves the heavy chain of factor X, producing the activated factor Xa alpha. The purified factor X activator from B. fasciatus venom did not activate prothrombin, nor did it cleave or clot purified fibrinogen. The amidolytic activity and the factor X activation activity of the factor X activator from B. fasciatus venom were readily inhibited by serine protease inhibitors such as diisopropyl fluorophosphate (DFP), phenylmethanesulfonyl fluoride (PMSF), benzamidine and by soybean trypsin inhibitor but not by EDTA. These observations suggest that the factor X activator from B. fasciatus venom is a serine protease. It therefore differs from those of activators obtained from Vipera russelli and Bothrops atrox venoms, which are metalloproteinases.

Characterization of a thrombin-like enzyme from the venom of Trimeresurus jerdonii.

From the venom of Trimeresurus jerdonii, a distinct thrombin-like enzyme, called jerdonobin, was purified by DEAE A-25 ion-exchange chromatography, Sephadex G-75 gel filtration, and fast protein liquid chromatography (FPLC). SDS-PAGE analysis of this enzyme shows that it consists of a single polypeptide chain with a molecular weight of 38,000. The NH(2)-terminal amino acid sequence of jerdonobin has great homology with venom thrombin-like enzymes documented. Jerdonobin is able to hydrolyze several chromogenic substrates. The enzyme directly clots fibrinogen with an activity of 217 NIH units/mg. The fibrinopeptides released, identified by HPLC, consisted of fibrinopeptide A and a small amount of fibrinopepide B. The activities of the enzyme were inhibited by phenylmethylsulfonyl fluoride (PMSF) and p-nitrophenyl-p-guanidinobenzoate (NPGB). However, metal chelator (EDTA) had no effect on it, indicating it is venom serine protease.

Isolation and properties of a blood coagulation factor X activator from the venom of king cobra (Ophiophagus hannah).

A specific blood coagulation factor X activator was purified from the venom of Ophiophagus hannah by gel filtration and two steps of FPLC Mono-Q column ion-exchange chromatography. It showed a single protein band both in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and alkaline polyacrylamide gel electrophoresis. The mol. wt was estimated to be 62,000 in non-reducing conditions and 64,500 in reducing conditions by SDS-PAGE. The isoelectric point was found to be pH 5.6. The enzyme had weak amidolytic activities toward CBS 65-25, but it showed no activities on S-2266, S-2302, thrombin substrate S-2238, plasmin substrate S-2251 or factor Xa substrate S-2222. It had no arginine esterase activity toward substrate benzoylarginine ethylester (BAEE). The enzyme activated factor X in vitro and the effect was absolutely Ca2+ dependent, with a Hill coefficient of 6.83. It could not activate prothrombin nor had any effect on fibrinogen and thus appeared to act specifically on factor X. The procoagulant activity of the enzyme was almost completely inhibited by serine protease inhibitors like PMSF, TPCK and soybean trypsin inhibitor; partially inhibited by L-cysteine. Metal chelator EDTA did not inhibit its procoagulant activity. These results suggest that the factor X activator from O. hannah venom is a serine protease.

Characterization of OhS1, an arginine/lysine amidase from the venom of king cobra (Ophiophagus hannah).

In this paper, we present the results of purification and characterization of an arginine/lysine amidase from the venom of Ophiophagus hannah (OhS1). It was purified by Sephadex G-75 gel filtration and ion-exchange chromatography on DEAE-Sepharose CL-6B. It is a protein of about 43,000, consisting of a single polypeptide chain. It is a minor component in the venom. The purified enzyme was capable of hydrolysing several tripeptidyl-p-nitroanilide substrates having either arginine or lysine as the C-terminal residue. We studied the kinetic parameters of OhS1 on six these chromogenic substrates. OhS1 did not clot fibrinogen. Electrophoresis of fibrinogen degraded with OhS1 revealed the disappearance of the alpha- and beta-chains and the appearance of lower mol. wt fragments. OhS1 had no hemorrhagic activity. It did not hydrolyse casein, nor did it act on blood coagulation factor X, prothrombin and plasminogen. The activity of OhS1 was completely inhibited by NPGB, PMSF, DFP, benzamidine and soybean trypsin inhibitor, suggesting it is a serine protease. Metal chelator (EDTA) had no effect on it.