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| 2. |
- Aleshkov, S B, et al.
(författare)
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Biochemical and biophysical studies of reactive center cleaved plasminogen activator inhibitor type 1. The distance between P3 and P1' determined by donor-donor fluorescence energy transfer.
- 1996
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Ingår i: Journal of Biological Chemistry. - 0021-9258 .- 1083-351X. ; 271:35, s. 21231-8
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Tidskriftsartikel (refereegranskat)abstract
- Plasminogen activator inhibitor type 1 (PAI-1) is a fast acting inhibitor of plasminogen activators (PAs). In accordance with other serpins, PAI-1 is thought to undergo a conformational change upon reactive center cleavage. In this study we have developed methods to produce and purify reactive center cleaved wild-type PAI-1 and characterized this molecular form of PAI-1 by biochemical and biophysical methods. Incubation with Sepharose-bound trypsin caused cleavage only at the P1-P1' bond in the reactive center and resulted in 39- and 4-kDa polypeptides, strongly held together by noncovalent interactions. Circular dichroism measurements suggest that the reactive center cleavage triggers larger conformational changes than the conversion from the active to the latent form. Cleaved PAI-1 did not bind to either PAs or vitronectin but retained the heparin-binding capacity. To study the structure of cleaved PAI-1 by polarized fluorescence spectroscopy and to measure intramolecular distances, we used cysteine substitution mutants to which extrinsic fluorescence probes were attached. These studies revealed increasing orientational freedom of probes in the P3 and P1' positions upon cleavage. Distance measurements based on fluorescence energy transfer between probes in positions P3 and P1' indicate that these residues are separated by at least 68 +/- 10 A in cleaved PAI-1.
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| 5. |
- GOOBARLARSSON, L, et al.
(författare)
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ENHANCEMENT OF HIV-1 PROTEINASE ACTIVITY BY HIV-1 REVERSE-TRANSCRIPTASE
- 1995
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Ingår i: VIROLOGY. - : ACADEMIC PRESS INC JNL-COMP SUBSCRIPTIONS. - 0042-6822. ; 206:1, s. 387-394
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- HIV-1 reverse transcriptase (RT) was found to increase the activity of HIV-I proteinase in vitro and in eukaryotic cells. The effect of RT on proteinase activity was dose-dependent and independent of pH or salt concentration. The cleavage of sequences cor
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| 7. |
- Annas, Anita, et al.
(författare)
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Basal and induced EROD activity in the chorioallantoic membrane during chicken embryo development
- 1999
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Ingår i: Environmental Toxicology and Pharmacology. - 1382-6689 .- 1872-7077. ; 8:1, s. 49-52
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- The chorioallantoic membrane (CAM) is a highly vascularized tissue that takes part in the respiratory exchange of gases through the eggshell. Although the CAM may be exposed to environmental contaminants, its response to pollutants has not been studied. We examined the cytochrome P4501A (CYP1A)-catalyzed deethylation of 7-ethoxyresorufin (EROD) in the CAM during chicken embryo development. EROD was constitutively present and was inducible by the aryl hydrocarbon (Ah) receptor agonist 3,3′,4,4′,5-pentachlorobiphenyl (PCB 126). Our results suggest the CAM as a first line of defence of the avian embryo against toxic compounds, but also as a target for CYP1A-activated chemicals.
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| 8. |
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| 9. |
- Fa, M, et al.
(författare)
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Time-resolved polarized fluorescence spectroscopy studies of plasminogen activator inhibitor type 1 : conformational changes of the reactive center upon interactions with target proteases, vitronectin and heparin.
- 1995
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Ingår i: Biochemistry. - 0006-2960 .- 1520-4995. ; 34:42, s. 13833-40
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Tidskriftsartikel (refereegranskat)abstract
- Plasminogen activator inhibitor type 1 (PAI-1) is an important physiological inhibitor of the plasminogen activator system. To investigate the structure-functional aspects of this inhibitor, we have taken advantage of the lack of cysteine residues in the PAI-1 molecule and substituted Ser344 (P3) and Met347 (P1'), in the reactive center loop, with cysteines, thereby creating unique attachment sites for extrinsic fluorescent probe. Both cysteine mutants were purified and labeled with a sulfhydryl specific fluorophore, N-(4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacen yl-3-propionyl)-N- (iodoacetyl)ethylenediamine (BDYIA). The labeled mutants were found to reveal biochemical characteristics very similar to those of wild type PAI-1. Time-resolved fluorescence spectroscopy was used to examine orientational freedom of BDYIA in the reactive center loop of PAI-1. The orientational freedom of the probe was found to be greater in the latent form than in the active form of PAI-1, suggesting that the reactive center has a more relaxed conformation in the latent form than in the active form. Complex formation with target proteases, tissue type plasminogen activator (tPA) and urokinase type plasminogen activator (uPA), caused decreased orientational freedom of BDYIA in the P3 position, while the orientational freedom of BDYIA in position P1' increased to a level similar to that of BDYIA in reactive center-cleaved PAI-1. In contrast, complex formation with modified anhydro-uPA, which is unable to cleave its substrate, largely restricted the orientational freedom of BDYIA probe in the P1' position.(ABSTRACT TRUNCATED AT 250 WORDS)
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| 12. |
- Joelson, T, et al.
(författare)
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Presentation of a foreign peptide on the surface of tomato bushy stunt virus
- 1997
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Ingår i: JOURNAL OF GENERAL VIROLOGY. - : SOC GENERAL MICROBIOLOGY. - 0022-1317. ; 78, s. 1213-1217
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- A 13-amino-acid peptide derived from the V3 loop of human immunodeficiency virus (HIV-1) glycoprotein 120 (gp120) was attached as a C-terminal gene fusion to the coat protein of tomato bushy stunt virus (TBSV). The architecture of this plant virus permitt
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