Mass Spectrometry versus Conventional Techniques of Protein Detection: Zika Virus NS3 Protease Activity towards Cellular Proteins.

Agnieszka Dabrowska,Joanna Ner-Kluza,Krzysztof Pyrc,Aleksandra Milewska,Piotr Suder

doi:10.3390/molecules26123732

Agnieszka Dabrowska, Joanna Ner-Kluza + Show 3 more

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https://doi.org/10.3390/molecules26123732

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Abstract

Mass spectrometry (MS) used in proteomic approaches is able to detect hundreds of proteins in a single assay. Although undeniable high analytical power of MS, data acquired sometimes lead to confusing results, especially during a search of very selective, unique interactions in complex biological matrices. Here, we would like to show an example of such confusing data, providing an extensive discussion on the observed phenomenon. Our investigations focus on the interaction between the Zika virus NS3 protease, which is essential for virus replication. This enzyme is known for helping to remodel the microenvironment of the infected cells. Several reports show that this protease can process cellular substrates and thereby modify cellular pathways that are important for the virus. Herein, we explored some of the targets of NS3, clearly shown by proteomic techniques, as processed during infection. Unfortunately, we could not confirm the biological relevance of protein targets for viral infections detected by MS. Thus, although mass spectrometry is highly sensitive and useful in many instances, also being able to show directions where cell/virus interaction occurs, we believe that deep recognition of their biological role is essential to receive complete insight into the investigated process.

Highlights

The NS2Bprotein and its inactive NS2B-NS3S135A variant were expressed from pBudCE4.1 plasmids in 293T and A549 cells
Western blotting with an anti-NS3 antibody detected NS2B-NS3WT and NS2B-NS3S135A in cell
The NS2BNS3 protein and its inactive NS2B-NS3S135A variant were expressed from pBudCE4.1 plasmids in 293T and A549 cells

Summary

Introduction

The Zika virus is a flavivirus discovered in 1947 in primates inhabiting the African. The virus was found to infect humans, for decades it was not considered to be a medical threat due to limited distribution and very mild symptoms associated with infection. More than 10 years ago, interest in the Zika virus began to increase as it became clear that the virus has broadened its geographic distribution, and the first outbreak was reported in the Federated States of Micronesia [2]. While the symptoms are relatively mild and include fever, rash, headache, and muscle pain, the infection may cause severe sequelae, and it is associated with Guillain–Barré syndrome [3]. The infection is most severe in pregnant women since it interferes with the development of the fetal cerebrum, predominantly resulting in microcephaly. A number of in vitro and in vivo studies have confirmed these observations [4,5,6,7,8,9,10]

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