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41.
Cloning, analysis, and bacterial expression of human farnesyl pyrophosphate synthetase and its regulation in Hep G2 cells 总被引:5,自引:0,他引:5
B T Sheares S S White D T Molowa K Chan V D Ding P A Kroon R G Bostedor J D Karkas 《Biochemistry》1989,28(20):8129-8135
A partial length cDNA encoding farnesyl pyrophosphate synthetase (hpt807) has been isolated from a human fetal liver cDNA library in lambda gt11. DNA sequence analysis reveals hpt807 is 1115 bp in length and contains an open reading frame coding for 346 amino acids before reaching a stop codon, a polyadenylation addition sequence, and the first 14 residues of a poly(A+) tail. Considerable nucleotide and deduced amino acid sequence homology is observed between hpt807 and previously isolated rat liver cDNAs for farnesyl pyrophosphate synthetase. Comparison with rat cDNAs suggests that hpt807 is about 20 bp short of encoding the initiator methionine of farnesyl pyrophosphate synthetase. The human cDNA was cloned into a prokaryotic expression vector and Escherichia coli strain DH5 alpha F'IQ was transformed. Clones were isolated that express an active fusion protein which can be readily observed on protein gels and specifically stained on immunoblots with an antibody raised against purified chicken farnesyl pyrophosphate phosphate synthetase. These data confirm the identity of hpt807 as encoding farnesyl pyrophosphate synthetase. Slot blot analyses of RNA isolated from Hep G2 cells show that the expression of farnesyl pyrophosphate synthetase mRNA is regulated. Lovastatin increases mRNA levels for farnesyl pyrophosphate synthetase 2.5-fold while mevalonic acid, low-density lipoprotein, and 25-hydroxycholesterol decrease mRNA levels to 40-50% of control values. 相似文献
42.
The primary structure of human apolipoprotein A-IV 总被引:2,自引:0,他引:2
C Y Yang Z W Gu I S Chong W J Xiong M Rosseneu H X Yang B R Lee A M Gotto L Chan 《Biochimica et biophysica acta》1989,1002(2):231-237
Human apolipoprotein (apo) A-IV was purified from chylous ascites fluid. Proteolytic peptides produced by trypsin and Staphylococcus aureus V8 proteinase digestions were purified by high-performance liquid chromatography and sequenced. Human apoA-IV contains 376 amino acid residues. The peptide-derived sequence generally matches two previously reported DNA-derived amino acid sequences except for discrepancies in five positions. In order to examine these discrepancies further, one complete apoA-IV cDNA clone and another partial clone were sequenced. Comparison of all the available information indicates that the peptide-derived sequence reported here is accurate. Sequencing errors probably account for some of the discrepancies between the two primary sequences predicted by earlier nucleotide analyses. In certain positions, however, bona fide sequence heterogeneity or cloning artifact cannot be excluded. 相似文献
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A 40 kilodalton rat liver nuclear protein binds specifically to apolipoprotein B mRNA around the RNA editing site. 总被引:8,自引:5,他引:3
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Apolipoprotein (apo) B-48 mRNA is the product of RNA editing which consists of a C----U conversion changing a CAA codon encoding Gln-2153 in apoB-100 mRNA to a UAA stop codon in apoB-48 mRNA. In the adult rat, RNA editing occurs both in the small intestine and the liver. We have studied the ability of rat liver nuclear extracts to bind to synthetic apoB mRNA segments spanning the editing site. Using an RNA gel mobility shift assay, we found the sequence-specific binding of a protein(s) to a 65-nucleotide apoB-100 mRNA. UV crosslinking followed by T1 ribonuclease digestion and SDS-polyacrylamide gel electrophoresis demonstrated the formation of a 40 kDa protein-RNA complex when 32P-labeled apoB-100 mRNA was incubated with a rat liver nuclear extract but not with HeLa nuclear extract. Binding was specific for the sense strand of apoB mRNA, and was not demonstrated with single-stranded apoB DNA, or antisense apoB RNA. The complex also failed to form if SDS was present during the UV light exposure. Binding experiments using synthetic apoB mRNAs indicate that the 40 kDa protein would also bind to apoB-48 mRNA but not apoA-I, apoA-IV, apoC-II or apoE mRNA. Experiments using deletion mutants of apoB-100 mRNA indicate efficient binding of wildtype 65-nucleotide (W65), 40-nucleotide (W40) and 26-nucleotide (W26) apoB-100 mRNA segments, but not 10-nucleotide (or smaller) segments of apoB-100 mRNA to the 40 kDa protein. In contrast, two other regions of apoB-100 mRNA, B-5' (bases 1128-3003) and B-3' (bases 11310-11390), failed to bind to the protein. The 40 kDa sequence-specific binding protein in rat liver nuclear extract may play a role in apoB-100 mRNA editing. 相似文献
47.
Inka Brockhausen Arthur A Grey Henrianna Pang Harry Schachter Jeremy P Carver 《Glycoconjugate journal》1988,5(4):419-448
Sixteen asparagine-linked oligosaccharides ranging in size from (Man)2(GlcNAc)2 (Fuc)1 to (GlcNAc)6(Man)3(GlcNAc)2 were obtained from human 1-acid glycoprotein and fibrinogen, hen ovomucoid and ovalbumin, and bovine fetuin, fibrin and thyroglobulin by hydrazinolysis, mild acid hydrolysis and glycosidase treatment. The oligosaccharides hadN-acetylglucosamine at the reducing termini and mannose andN-acetylglucosamine residues at the non-reducing termini and were prepared for use asN-acetylglucosaminyltransferase substrates. Purification of the oligosaccharides involved gel filtration and high performance liquid chromatography on reverse phase and amine-bonded silica columns. Structures were determined by 360 MHz and 500 MHz proton nuclear magnetic resonance spectroscopy, fast atom bombardment-mass spectrometry and methylation analysis. Several of these oligosaccharides have not previously been well characterized.Abbreviations bis
bisecting GlcNAc
- DMSO
dimethylsulfoxide
- FAB
fast atom bombardment
- Fuc
l-fucose
- Gal
d-galactose
- GLC
gas-liquid chromatography
- GlcNAc or Gn
N-acetyl-d-glucosamine
- HPLC
high performance liquid chromatography
- Man or M
d-mannose
- MES
2-(N-morpholino)ethanesulfonate
- MS
mass spectrometry
- NMR
nuclear magnetic resonance
- PIPES
piperazine-N,N-bis(2-ethane sulfonic acid)
the nomenclature of the oligosaccharides is shown in Table 1. 相似文献
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Two analogues of angiotensin III were compared as antagonists of the pressor response to angiotensin II (ANG II) and angiotensin III (ANG III) in conscious, unrestrained rats. Dose-mean arterial pressure (MAP) response curves were obtained for ANG II and ANG III in the absence or presence of [Ile7]ANG III (1.3 x 10(-7) mol/kg) or [Sar1 Ile7]ANG III (1.2 x 10(-7) mol/kg). In the presence of [Ile7]ANG III, the dose-MAP response curves for ANG II and ANG III were significantly displaced to the right. [Ile7]ANG III behaved as a partial agonist on ANG II but not ANG III receptors. In the presence of [Sar1 Ile7]ANG III, the dose-MAP response curve for ANG III but not ANG II was significantly displaced to the right. This suggests that [Sar1 Ile7]ANG III is a selective antagonist of ANG III in the vasculature. [Ile7]ANG III, on the other hand, antagonizes both ANG II and ANG III receptors. Our results support the hypothesis of the existence of a sub-class of angiotensin receptors activated by ANG III in the vascular smooth muscle. 相似文献