TY - JOUR
T1 - Self-assembling functional programmable protein array for studying protein–protein interactions in malaria parasites
AU - Arévalo-Pinzón, Gabriela
AU - González-González, María
AU - Suárez, Carlos Fernando
AU - Curtidor, Hernando
AU - Carabias-Sánchez, Javier
AU - Muro, Antonio
AU - LaBaer, Joshua
AU - Patarroyo, Manuel Alfonso
AU - Fuentes, Manuel
N1 - Funding Information:
This research was financed by the Colombian Departamento Administra‑ tivo de Ciencia, Tecnología e Innovación (COLCIENCIAS) through contract RC# 0309‑2013 and the Spanish Institute of Health Carlos III (ISCIII) through Grant FIS PI 14/01538 & FIS PI 17/01930. We would also like to acknowledge contributions from the European Regional Development Fund (ERDF), the Junta Castilla‑León (BIO/SA07/15) and Fundación Solórzano (FS‑23‑2015). The Proteomics Unit forms part of ProteoRed, PRB3‑ISCIII, supported by Grant PT17/0019/0023, PE I+D+I 2017‑2012, funded by ISCIII and ERDF. The Proteomics Unit belongs to ProteoRed, PRB3‑ISCIII, supported by Grant PT17/0019/0023. The work was carried out during Arévalo‑Pinzón’s PhD internship financed by COLCIENCIAS within the framework of the “Convoca‑ toria Nacional para Estudios de Doctorado en Colombia (call for candidates no. 567)”.
Publisher Copyright:
© 2018 The Author(s).
Copyright:
Copyright 2018 Elsevier B.V., All rights reserved.
PY - 2018/7/17
Y1 - 2018/7/17
N2 - Background: Plasmodium vivax is the most widespread malarial species, causing significant morbidity worldwide. Knowledge is limited regarding the molecular mechanism of invasion due to the lack of a continuous in vitro culture system for these species. Since protein-protein and host-cell interactions play an essential role in the microorganism’s invasion and replication, elucidating protein function during invasion is critical when developing more effective control methods. Nucleic acid programmable protein array (NAPPA) has thus become a suitable technology for studying protein-protein and host-protein interactions since producing proteins through the in vitro transcription/translation (IVTT) method overcomes most of the drawbacks encountered to date, such as heterologous protein production, stability and purification. Results: Twenty P. vivax proteins on merozoite surface or in secretory organelles were selected and successfully cloned using gateway technology. Most constructs were displayed in the array expressed in situ, using the IVTT method. The Pv12 protein was used as bait for evaluating array functionality and co-expressed with P. vivax cDNA display in the array. It was found that Pv12 interacted with Pv41 (as previously described), as well as PvMSP142kDa, PvRBP1a, PvMSP8 and PvRAP1. Conclusions: NAPPA is a high-performance technique enabling co-expression of bait and query in situ, thereby enabling interactions to be analysed rapidly and reproducibly. It offers a fresh alternative for studying protein-protein and ligand-receptor interactions regarding a parasite which is difficult to cultivate (i.e. P. vivax).
AB - Background: Plasmodium vivax is the most widespread malarial species, causing significant morbidity worldwide. Knowledge is limited regarding the molecular mechanism of invasion due to the lack of a continuous in vitro culture system for these species. Since protein-protein and host-cell interactions play an essential role in the microorganism’s invasion and replication, elucidating protein function during invasion is critical when developing more effective control methods. Nucleic acid programmable protein array (NAPPA) has thus become a suitable technology for studying protein-protein and host-protein interactions since producing proteins through the in vitro transcription/translation (IVTT) method overcomes most of the drawbacks encountered to date, such as heterologous protein production, stability and purification. Results: Twenty P. vivax proteins on merozoite surface or in secretory organelles were selected and successfully cloned using gateway technology. Most constructs were displayed in the array expressed in situ, using the IVTT method. The Pv12 protein was used as bait for evaluating array functionality and co-expressed with P. vivax cDNA display in the array. It was found that Pv12 interacted with Pv41 (as previously described), as well as PvMSP142kDa, PvRBP1a, PvMSP8 and PvRAP1. Conclusions: NAPPA is a high-performance technique enabling co-expression of bait and query in situ, thereby enabling interactions to be analysed rapidly and reproducibly. It offers a fresh alternative for studying protein-protein and ligand-receptor interactions regarding a parasite which is difficult to cultivate (i.e. P. vivax).
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U2 - 10.1186/s12936-018-2414-2
DO - 10.1186/s12936-018-2414-2
M3 - Research Article
C2 - 30016987
AN - SCOPUS:85050126715
SN - 1475-2875
VL - 17
JO - Malaria Journal
JF - Malaria Journal
IS - 1
M1 - 270
ER -