Biochemical characterization of naturally occurring mutations in SARS-CoV-2 RNA-dependent RNA polymerase

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F60461373%3A22330%2F24%3A43930726" target="_blank" >RIV/60461373:22330/24:43930726 - isvavai.cz</a>

Nalezeny alternativní kódy

RIV/60461373:22810/24:43930726

Výsledek na webu

<a href="https://onlinelibrary.wiley.com/doi/10.1002/pro.5103" target="_blank" >https://onlinelibrary.wiley.com/doi/10.1002/pro.5103</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1002/pro.5103" target="_blank" >10.1002/pro.5103</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Biochemical characterization of naturally occurring mutations in SARS-CoV-2 RNA-dependent RNA polymerase

Popis výsledku v původním jazyce

Since the emergence of SARS-CoV-2, mutations in all subunits of the RNA-dependent RNA polymerase (RdRp) of the virus have been repeatedly reported. Although RdRp represents a primary target for antiviral drugs, experimental studies exploring the phenotypic effect of these mutations have been limited. This study focuses on the phenotypic effects of substitutions in the three RdRp subunits: nsp7, nsp8, and nsp12, selected based on their occurrence rate and potential impact. We employed nano-differential scanning fluorimetry and microscale thermophoresis to examine the impact of these mutations on protein stability and RdRp complex assembly. We observed diverse impacts; notably, a single mutation in nsp8 significantly increased its stability as evidenced by a 13 degrees C increase in melting temperature, whereas certain mutations in nsp7 and nsp8 reduced their binding affinity to nsp12 during RdRp complex formation. Using a fluorometric enzymatic assay, we assessed the overall effect on RNA polymerase activity. We found that most of the examined mutations altered the polymerase activity, often as a direct result of changes in stability or affinity to the other components of the RdRp complex. Intriguingly, a combination of nsp8 A21V and nsp12 P323L mutations resulted in a 50% increase in polymerase activity. To our knowledge, this is the first biochemical study to demonstrate the impact of amino acid mutations across all components constituting the RdRp complex in emerging SARS-CoV-2 subvariants.

Název v anglickém jazyce

Biochemical characterization of naturally occurring mutations in SARS-CoV-2 RNA-dependent RNA polymerase

Popis výsledku anglicky

Since the emergence of SARS-CoV-2, mutations in all subunits of the RNA-dependent RNA polymerase (RdRp) of the virus have been repeatedly reported. Although RdRp represents a primary target for antiviral drugs, experimental studies exploring the phenotypic effect of these mutations have been limited. This study focuses on the phenotypic effects of substitutions in the three RdRp subunits: nsp7, nsp8, and nsp12, selected based on their occurrence rate and potential impact. We employed nano-differential scanning fluorimetry and microscale thermophoresis to examine the impact of these mutations on protein stability and RdRp complex assembly. We observed diverse impacts; notably, a single mutation in nsp8 significantly increased its stability as evidenced by a 13 degrees C increase in melting temperature, whereas certain mutations in nsp7 and nsp8 reduced their binding affinity to nsp12 during RdRp complex formation. Using a fluorometric enzymatic assay, we assessed the overall effect on RNA polymerase activity. We found that most of the examined mutations altered the polymerase activity, often as a direct result of changes in stability or affinity to the other components of the RdRp complex. Intriguingly, a combination of nsp8 A21V and nsp12 P323L mutations resulted in a 50% increase in polymerase activity. To our knowledge, this is the first biochemical study to demonstrate the impact of amino acid mutations across all components constituting the RdRp complex in emerging SARS-CoV-2 subvariants.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10608 - Biochemistry and molecular biology

Projekt

Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.

Návaznosti

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Rok uplatnění

2024

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

PROTEIN SCIENCE

ISSN

0961-8368

e-ISSN

1469-896X

Svazek periodika

33

Číslo periodika v rámci svazku

9

Stát vydavatele periodika

ZA - Jihoafrická republika

Počet stran výsledku

16

Strana od-do

—

Kód UT WoS článku

001291756000001

EID výsledku v databázi Scopus

2-s2.0-85201356758