Purification and characterization of camel liver L-asparaginase

L-asparaginase from camel liver was isolated and purified by heat denaturation followed by QAE-Sephadex A-50 column chromatography and SP-Sepharose column chromatography. The purified camel liver L-asparaginase had a molecular weight of 180 kDa [consistent with a homotetramer] and a pI value of 8.6. The enzyme was thermostable with relative structure rigidity and an optimum temperature at 65°C. It had a pH optimum at 9.6 and was stable for storage at 4°C in the refrigerator for 7 days

Expression of recombinant escherichia coli L-asparaginase II, purification and characterization

L-Asparaginase [ASNase] is an anti-cancer [[antineoplastic] or [cytotoxic]] chemotherapy drug that is used for the treatment of acute lymphoblastic leukemia [ALL]. An efficient and economical scheme was developed for over expression and rapid purification of the Escherichia coli enzyme. The gene encoding for the Escherichia coli L-asparaginase was PCR-amplified and cloned in pGEX-4Tl expression vector. The recombinant L-asparaginase was purified to homogeneity by affinity chromatography on glutathione Sepharose column. The recombinant enzyme had an apparent MW of 152 kDa and a K[m] value of 12.5 microM for the main physiological substrate L-asparagine. The pI value was 5.6 while the turnover number [catalytic constant] was 1 x 10[2] s[-1] and the K[cat]/K[m] value [specificity constant] was 0.8 x 10[7] M[-1]s[-1]

Matrix metalloproteinase-1 in camel liver tissues: isolation, purification and some characterization

Matrix metalloproteinases [MMPs] constitute a multigene family of over 25 secreted and cell surface enzymes that process or degrade numerous pericellular substrates. Their targets include other proteinases, clotting factors, cell surface receptors, and virtually all structural extracellular matrix proteins The MMPs play a vital role in cellular functions as cell proliferation, tissue development, wound healing and tissue repair. In this study, a trial was carried out to isolate matrix metalloproteinase-l [MMP-1: collagenase, EC 3.4.24.7] from camel liver tissues [CaInelus dromedarius] in a small scale purification technique using ammonium sulphate fractionation, gel filtration on Sephadex G-100 and Bio-Gel A-1.5m column chromatographies. The homogeneity of the purified preparation was judged by polyacrylamide gel electrophoresis [PAGE] in the presence and absence of SDS and beta-mercaptoethanol. The enzyme is heterodimer and the molecular weight [M[r]] of the two subunits are 67 and 30 kDa, while that of the native enzyme is 97 kDa. The optimum temperature was in the range of 30-40°C EDTA was more potent inhibitor than DTT for enzyme activity

Role of mismatch repair genes [h MLHI, h PMS1, h PMS2, GTBP/h MSH6, h MSH2] in the pathogenesis of hepatocellular carcinoma

DNA mismatch repair [MMR] is an important mechanism involved in maintaining fidelity of genomic DNA. Abnormalities in at least one of five MMR genes are implicated in the development of many cancers and the associated micro satellite instability [MSI]. By using a newly developed multiplex reverse transcription -PCR assay, the expression of the five known MMR [hMLH1, hPMS1, hPMS2, GTBP/hMSH6, hMSH2] were evaluated in 33 human HCC cases as well as 16 cases from the normal distant hepatic tissue samples [NDHT] were also evaluated. Twenty- five of them were associated with HCV infection. This was done in an attempt to determine the role of MMR genes in the development of HCC. The beta actin gene was used as an internal control for RNA degradation and DNA contamination and as well as a reference for quantifying the levels of their transcription. Out of the 33 studied HCC cases, 30 cases [90.9%] showed reduction in the expression of one or more of the 5 studied MMR genes. Reduced expression of hMSH2 was found in [71.9%], hMLH1 [53.3%], GTBP [51.1%], hPMS2 [33.3%] and hPMSI [6%]. Correlation analysis showed a strong significant correlation [P= 0.0069] between reduced expression of hPMS2 and GTBP [P=0.0034] as well as hPMS2 and non-cirrhosis [P=0.0197]. Chi-square analysis showed a significant correlation between reduced expression of hMLHl and grade II. On the other hand, 57.1%, 50%, 20%, 18.8% and 6% of the NDHT showed reduced expression of hMSH2, hMLHI, GTBP, hPMS2 and hPMSI respectively. Multivariate analysis showed significant correlation between HCC and hMSH2 [P= 0.008], hMLH1 [P=0.001] and GTBP [P=0.032], also between hPMS2, GTBP and HCC infected with HCV cases [P< 0.001, 0.002]. It could finally be concluded that reduced expression of hPMS2 is likely associated with growth advantage and stimulates proliferation changes that have encouraged malignant development in non- cirrhotic HCV infected patients via acquisition of more genetic damage and the MMR defects that occur at an early stage of hepatocarcinogenesis

Urinary thromboxane B2: a new predicting marker for acute myocardial infarction in subjects with risk factors

The authors evaluated the possible significant role of urinary TXB2 as a new predicting marker of Acute Myocardial Infarction [AMI] in high-risk subjects. Eighty persons were submitted to this study and divided into three groups, the control group [n = 20], the high risk group [n = 30] and the AMI group [n =30]. all persons subjected to clinical and laboratory evaluation: Cardiac enzymes, Myoglobin, Lipid profiles, Pyruvate Kinase and urinary TXB2 level. A high significant elevation of Cardiac enzymes, Myoglobin and Pyruvate Kinase was observed in AMI group only. Urinary TXB2 in risk group was highly significant elevated while in AMI group was non significant increased compared to control group the Urinary TXB2 could be used as a good predicting marker for probability of AMI in patients who had one or more of the AMI risk factors