Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.23 (beta-galactosidase)
 14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have developed a sandwich-enzyme immunoassay (EIA) for the quantification of lipoprotein lipase (LPL) and hepatic triglyceride lipase (HTGL) in human postheparin plasma (PHP) using monoclonal antibodies (MAbs) directed against the corresponding enzymes purified from human PHP. The sandwich-EIA for LPL was performed by using the combination of two distinct types of anti-LPL MAbs that recognize different epitopes on the LPL molecule. The immunoreactive mass of LPL was specifically measured using a beta-galactosidase-labeled anti-LPL MAb as an enzyme-linked MAb, an anti-LPL MAb linked with the bacterial cell wall as an insolubilized MAb, and purified human PHP-LPL as a standard. The sandwich-EIA for HTGL was carried out by using two distinct anti-HTGL MAbs that recognize different epitopes on HTGL. The limit of detection was 20 ng/ml for LPL and 60 ng/ml for HTGL. Each method yielded a coefficient of variation of less than 6% in intra- and inter-assays, and a high concentration of triglyceride did not interfere with the assays. The average recovery of purified human PHP-LPL and -HTGL added to human PHP samples was 98.8% and 97.5%, respectively. The immunoreactive masses of LPL and HTGL in PHP samples, obtained at a heparin dose of 30 IU/kg, from 34 normolipidemic and 20 hypertriglyceridemic subjects were quantified by the sandwich-EIA. To assess the reliability of the measured mass values, they were compared with the corresponding enzyme activities measured by selective immunoinactivation assay using rabbit anti-human PHP-LPL and -HTGL polyclonal antisera. Both assay methods yielded a highly significant correlation in either normolipidemic (r = 0.945 for LPL; r = 0.932 for HTGL) or hypertriglyceridemic subjects (r = 0.989 for LPL; r = 0.954 for HTGL). The normal mean (+/- SD) level of lipoprotein lipase mass and activity in postheparin plasma was 223 +/- 66 ng/ml and 10.1 +/- 2.9 mumol/h per ml, and that of hepatic triglyceride lipase mass and activity was 1456 +/- 469 ng/ml and 26.4 +/- 8.7 mumol/h per ml, respectively. The present sandwich-enzyme immunoassay methods make it possible to study the molecular nature of LPL and HTGL in PHP from patients with either primary or secondary hyperlipoproteinemia.
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PMID:A sandwich-enzyme immunoassay for the quantification of lipoprotein lipase and hepatic triglyceride lipase in human postheparin plasma using monoclonal antibodies to the corresponding enzymes. 207 12

A full-length human apo CII cDNA clone has been constructed by completing the 5' end of an incomplete cDNA with a 44 bp long synthetic oligonucleotide. This apo CII cDNA insert was cloned into the pSP19 expression vector and transcribed and translated in vitro. Its N-terminal signal sequence (23 amino-acid residues) was accurately cleaved during cotranslational translocation through endoplasmic reticulum membranes to yield the mature apo CII. Mature apo CII was expressed on a preparative scale as fusion protein apo CII-beta-galactosidase with the full-length apo CII cDNA integrated into the pUR291 vector. Furthermore it was expressed in E. coli transformed with the pKK233-2 apo CII clone. The preform was accurately processed by the host cell. C-Terminal apo CII deletion mutants generated by partial Bal31 digestion of the pKK233-2 apo CII vector yielded well-defined truncated apo CII polypeptides on a preparative scale which allowed the determination of the polypeptide domain responsible for the activation of the serum lipoprotein lipase.
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PMID:Expression of normal and mutagenized apolipoprotein CII in procaryotic cells. Structure-function relationship. 306 32

To increase the number of genes that can be mapped to the genome of the tammar wallaby (Macropus eugenii), we sequenced 100 randomly chosen clones from a mammary gland cDNA library. Provisional identifications were made of seven nuclear genes and one mitochondrial gene encoding two caseins, beta-galactosidase, acetyl-coenzyme A synthetase, lipoprotein lipase, inorganic pyrophosphatase, an ATP-dependent RNA helicase, and cytochrome c oxidase I. Highly conserved genes, such as that encoding acetyl-coenzyme A synthetase, were easily identified even from cross-kingdom matches. Genes which are highly divergent, however, such as those encoding the mature casein peptides, could not be aligned with homologues in the databases. Even in an organ where there is high mRNA species redundancy, the sequence characterization of expressed sequence tags provides a rapid means of gene identification for mapping purposes.
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PMID:The identification of nuclear and mitochondrial genes by sequencing randomly chosen clones from a marsupial mammary gland cDNA library. 799 73

Gene therapy to deliver and express a corrective lipoprotein lipase (LPL) gene may improve the lipid profile and reduce the morbidity and potential atherogenic risk from hypertriglyceridemia and dyslipoproteinemia in patients with complete or partial LPL deficiency. We have used an E1-/E3- adenoviral vector, with an RSV-driven human LPL cDNA expression cassette (Ad-RSV-LPL), to achieve high ectopic LPL gene expression in the human hepatoma cell line HepG2, an accepted hepatocellular model of lipoprotein metabolism. Ad-RSV-LPL transduction of HepG2 cells with a multiplicity of infection (moi) between 12.5 and 100 yielded dose-dependent increments in LPL mass and activity. Peak levels of LPL protein of 2,032.1 +/- 274.5 ng/10(5) cells per ml (mol 100) correlated with increased activity of 92.7 +/- 22.6 mU/10(5) cells per ml relative to negligible LPL levels in Ad-RSV-LacZ (beta-galactosidase) controls. Exogenous LPL expression over a 5-day period peaked at day 3. Susceptibility to inhibition by 1 M NaCl and an anti-LPL monoclonal antibody confirmed that lipase activity was indeed derived from human LPL. Hydrolysis, by LPL-overexpressing HepG2 cells, of TG carried in very-low-density lipoprotein (VLDL) showed that greater than 50% of the triglycerides (TG) disappeared after 4 hr of incubation. These results were compatible with FPLC evidence of a marked reduction in VLDL-TG. These results provide strong in vitro evidence that adenoviral-mediated ectopic expression of the human LPL gene could render hepatic cells capable of VLDL catabolism and thus support the possibility for in vivo adenoviral vector-mediated liver-targeted LPL gene therapy.
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PMID:Efficient adenovirus-mediated ectopic gene expression of human lipoprotein lipase in human hepatic (HepG2) cells. 901 24

Humans homozygous or heterozygous for mutations in the lipoprotein lipase (LPL) gene demonstrate significant disturbances in plasma lipoproteins, including raised triglyceride (TG) and reduced HDL cholesterol levels. In this study we explored the feasibility of adenovirus-mediated gene replacement therapy for LPL deficiency. A total of 5 x 10(9) plaque-forming units (pfu) of an E1/E3-deleted adenovirus expressing either human LPL (Ad-LPL) or the bacterial beta-galactosidase gene (Ad-LacZ) as a control were administered to mice heterozygous for targeted disruption in the LPL gene (n = 57). Peak expression of total postheparin plasma LPL activity was observed at day 7 in Ad-LPL mice versus Ad-LacZ controls (834 +/- 133 vs 313 +/- 89 mU/mL, P < .01), and correlated with human-specific LPL activity (522 +/- 219 mU/mL) and mass (9214 +/- 782 ng/mL), a change that was significant to 14 and 42 days, respectively. At day 7, plasma TGs were significantly reduced relative to Ad-LacZ mice (0.17 +/- 0.07 vs 1.90 +/- 0.89 mmol/L, P < .01) but returned to endogenous levels by day 42. Ectopic liver expression of human LPL was confirmed by in situ hybridization analysis and from raised LPL activity and mass in liver homogenates. Analysis of plasma lipoprotein composition revealed a marked decrease in VLDL-derived TGs. Severely impaired oral and intravenous fat-load tolerance in LPL-deficient mice was subsequently corrected after Ad-LPL administration and closely paralleled that observed in wild-type mice. These findings suggest that liver-targeted adenovirus-mediated LPL gene transfer offers an effective means for transient correction of altered lipoprotein metabolism and impaired fat tolerance due to LPL deficiency.
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PMID:Correction of hypertriglyceridemia and impaired fat tolerance in lipoprotein lipase-deficient mice by adenovirus-mediated expression of human lipoprotein lipase. 940 24

Much of the knowledge about the cell biology of lipoprotein lipase (LPL) in vitro has been gained from adipose tissue model systems. However, the importance of skeletal muscle lipoprotein lipase (SMLPL) to both lipoprotein and muscle metabolism remains unclear. Although the production of LPL in cultured myocytes has been documented, the amount of enzyme activity produced is small. To develop a more suitable tissue culture model for SMLPL, mouse C(2)C(12) myoblasts were stably transduced with a retroviral vector encoding the full-length human LPL (hLPL) cDNA. Control cells were transduced with a vector encoding beta-galactosidase. LPL expression was assayed as a function of cell growth by measuring LPL activity on days 3, 7, 9, 11, and 14 after subculture. The hLPL-transduced myoblasts increasingly overexpressed both heparin-releasable (HR) and intracellular (IN) LPL activity compared to nontransduced myoblasts (P < 0.001 at Day 11) and myoblasts transduced with the control vector (P < 0.001 at Day 11). This increase occurred while LPL mRNA levels remained stable between days 3 and 14. As expected, IN LPL activity was also increased in the transduced cells. High levels of LPL activity were also obtained after differentiating the C(2)C(12) cells into myotubes by serum deprivation. Additionally, throughout the time course, C(2)/LPL cells had greater amounts of intracellular triglyceride than both the C(2)C(12) and the C(2)/beta-GEO cells (P = 0.005 and P < 0.001, respectively) with the largest differences seen on day 14 of the time course (P = 0.001, C(2)/LPL vs C(2)C(12) (r) or C(2)/beta-GEO cells). Thus, C(2)C(12) myoblasts stably transduced with hLPL markedly overexpressed both HR and IN LPL activity compared to control cells which, in turn, was associated with increases in intracellular triglyceride content. Because LPL regulation in tissues is mostly posttranslational, this new in vitro model will permit the in-depth study of the posttranslational regulation of SMLPL and provide new insights into the fate of lipoprotein-derived fatty acids in muscle.
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PMID:Increased intracellular triglyceride in C(2)C(12) muscle cells transfected with human lipoprotein lipase. 1077 40