Exploring the application of ELISA and slot blot as utility protein–protein interaction analysis tools

Abstract

Although protein interaction studies are instrumental in understanding protein networks, most protein interaction techniques depend on use of sophisticated and expensive equipment. This makes it difficult for under-resourced laboratories to conduct protein–protein interaction studies. As such, we sought to explore the prospects of using ELISA and slot blot as alternate methods for analyzing protein–protein interactions in resource-limited settings. We used these two methods to explore the well established interaction of heat shock proteins (Hsps) of Plasmodium falciparum as a model. P. falciparum Hsp70-1 (PfHsp70-1) is a cytosol-nuclear localised molecular chaperone that interacts with several functional partners including P. falciparum Hsp70-Hsp90 organising protein (PfHop), P. falciparum Hsp40 (PfHsp40) and P. falciparum Hsp70-z (PfHsp70-z). To validate the application of ELISA and slot blot techniques in protein–protein studies, we employed these two techniques to explore the interaction of recombinant PfHsp70-1 with its partners. We further used the two techniques to explore the effects of mutating residues located in the GGMP repeat and linker motifs of PfHsp70-1 on the chaperone’s interaction with its functional partners. We established that despite requiring much larger amounts of protein compared to the more sensitive assays, the ELISA and slot blot assays were capable of detecting both nucleotide- and mutation-driven changes regulating the affinity of PfHsp70-1 for its interactors. Our findings highlight the utility of these two techniques under resource constraints in conducting routine protein–protein interaction studies and their possible application in the preliminary screening of inhibitors targeting protein networks.

KEYWORDS:

• protein–protein interaction
• heat shock protein
• slot blot assay
• ELISA
• SPR
• inhibitor screening

ACKNOWLEDGEMENTS

The authors would like to acknowledge the University of Cape Town, Faculty of Health Sciences for financial support to SM and Stellenbosch University Sub-Committee B for financial support to TZ.

DISCLOSURE

The authors declare no potential conflict of interest.

SUPPLEMENTAL DATA

Supplemental data for this article can be accessed here https://doi.org/10.1080/0035919X.2022.2144536

Additional information

Funding

The study was supported through a Digital Cooperation Fellowship awarded to AS by the Alexander von Humboldt Foundation of Germany. This work is based on the research supported in part by the Department of Science and Technology/National Research Foundation (NRF) of South Africa (Grant numbers, 75464 & 92598) awarded to A.S; and (Grant numbers 129401 & 145405) awarded to TZ.