Protein-binding RNA aptamers affect molecular interactions distantly from their binding sites.
(2015) In PLoS ONE 10(3).- Abstract
- Nucleic acid aptamer selection is a powerful strategy for the development of regulatory agents for molecular intervention. Accordingly, aptamers have proven their diligence in the intervention with serine protease activities, which play important roles in physiology and pathophysiology. Nonetheless, there are only a few studies on the molecular basis underlying aptamer-protease interactions and the associated mechanisms of inhibition. In the present study, we use site-directed mutagenesis to delineate the binding sites of two 2´-fluoropyrimidine RNA aptamers (upanap-12 and upanap-126) with therapeutic potential, both binding to the serine protease urokinase-type plasminogen activator (uPA). We determine the subsequent impact of aptamer... (More)
- Nucleic acid aptamer selection is a powerful strategy for the development of regulatory agents for molecular intervention. Accordingly, aptamers have proven their diligence in the intervention with serine protease activities, which play important roles in physiology and pathophysiology. Nonetheless, there are only a few studies on the molecular basis underlying aptamer-protease interactions and the associated mechanisms of inhibition. In the present study, we use site-directed mutagenesis to delineate the binding sites of two 2´-fluoropyrimidine RNA aptamers (upanap-12 and upanap-126) with therapeutic potential, both binding to the serine protease urokinase-type plasminogen activator (uPA). We determine the subsequent impact of aptamer binding on the well-established molecular interactions (plasmin, PAI-1, uPAR, and LRP-1A) controlling uPA activities. One of the aptamers (upanap-126) binds to the area around the C-terminal α-helix in pro-uPA, while the other aptamer (upanap-12) binds to both the β-hairpin of the growth factor domain and the kringle domain of uPA. Based on the mapping studies, combined with data from small-angle X-ray scattering analysis, we construct a model for the upanap-12:pro-uPA complex. The results suggest and highlight that the size and shape of an aptamer as well as the domain organization of a multi-domain protein such as uPA, may provide the basis for extensive sterical interference with protein ligand interactions considered distant from the aptamer binding site. (Less)
Please use this url to cite or link to this publication:
https://lup.lub.lu.se/record/5258007
- author
- Dupont, Daniel M ; Thuesen, Cathrine K ; Bøtkjær, Kenneth A ; Behrens, Manja LU ; Dam, Karen ; Sørensen, Hans P ; Pedersen, Jan S ; Ploug, Michael ; Jensen, Jan K and Andreasen, Peter A
- organization
- publishing date
- 2015
- type
- Contribution to journal
- publication status
- published
- subject
- in
- PLoS ONE
- volume
- 10
- issue
- 3
- article number
- e0119207
- publisher
- Public Library of Science (PLoS)
- external identifiers
-
- pmid:25793507
- wos:000352084200061
- scopus:84925949729
- pmid:25793507
- ISSN
- 1932-6203
- DOI
- 10.1371/journal.pone.0119207
- language
- English
- LU publication?
- yes
- id
- 3283633e-3aae-4f8a-970b-fc4d4cc5d6ae (old id 5258007)
- date added to LUP
- 2016-04-01 13:11:06
- date last changed
- 2022-03-13 22:40:02
@article{3283633e-3aae-4f8a-970b-fc4d4cc5d6ae,
abstract = {{Nucleic acid aptamer selection is a powerful strategy for the development of regulatory agents for molecular intervention. Accordingly, aptamers have proven their diligence in the intervention with serine protease activities, which play important roles in physiology and pathophysiology. Nonetheless, there are only a few studies on the molecular basis underlying aptamer-protease interactions and the associated mechanisms of inhibition. In the present study, we use site-directed mutagenesis to delineate the binding sites of two 2´-fluoropyrimidine RNA aptamers (upanap-12 and upanap-126) with therapeutic potential, both binding to the serine protease urokinase-type plasminogen activator (uPA). We determine the subsequent impact of aptamer binding on the well-established molecular interactions (plasmin, PAI-1, uPAR, and LRP-1A) controlling uPA activities. One of the aptamers (upanap-126) binds to the area around the C-terminal α-helix in pro-uPA, while the other aptamer (upanap-12) binds to both the β-hairpin of the growth factor domain and the kringle domain of uPA. Based on the mapping studies, combined with data from small-angle X-ray scattering analysis, we construct a model for the upanap-12:pro-uPA complex. The results suggest and highlight that the size and shape of an aptamer as well as the domain organization of a multi-domain protein such as uPA, may provide the basis for extensive sterical interference with protein ligand interactions considered distant from the aptamer binding site.}},
author = {{Dupont, Daniel M and Thuesen, Cathrine K and Bøtkjær, Kenneth A and Behrens, Manja and Dam, Karen and Sørensen, Hans P and Pedersen, Jan S and Ploug, Michael and Jensen, Jan K and Andreasen, Peter A}},
issn = {{1932-6203}},
language = {{eng}},
number = {{3}},
publisher = {{Public Library of Science (PLoS)}},
series = {{PLoS ONE}},
title = {{Protein-binding RNA aptamers affect molecular interactions distantly from their binding sites.}},
url = {{http://dx.doi.org/10.1371/journal.pone.0119207}},
doi = {{10.1371/journal.pone.0119207}},
volume = {{10}},
year = {{2015}},
}