Cofactor induced dissociation of the multifunctional multisubunit EcoR124I investigated using electromobility shift assays, AFM and SPR

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F61388971%3A_____%2F17%3A00477805" target="_blank" >RIV/61388971:_____/17:00477805 - isvavai.cz</a>

Výsledek na webu

<a href="http://dx.doi.org/10.1039/c7ra07505g" target="_blank" >http://dx.doi.org/10.1039/c7ra07505g</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1039/c7ra07505g" target="_blank" >10.1039/c7ra07505g</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Cofactor induced dissociation of the multifunctional multisubunit EcoR124I investigated using electromobility shift assays, AFM and SPR

Popis výsledku v původním jazyce

We have applied three techniques to the study of subunit assembly of the Type IC Restriction-Modification enzyme EcoR124I. This fully functional enzyme EcoR124I consists of a complex of the three subunits HsdR, HsdM and HsdS in a R2M2S1 stoichiometry, but is known to dissociate readily, releasing free HsdR and producing first an R-1-complex and then the core, DNA-binding methyltransferase (M2S1) complex. Analysis of the assembly pathway of this enzyme has previously employed gel retardation and Surface Plasmon Resonance (SPR), but the studies to date have not included the cofactors required for full enzyme activity. In this paper, we have also used atomic force microscopy (AFM)-based molecular volume measurements, and have analysed the effect of the cofactors ATP and AdoMet on enzyme stability and subunit assembly. We compare the data obtained from all three techniques and we show that they all give consistent results, but inherent differences in the methodologies provide additional information useful for the study of subunit assembly.

Název v anglickém jazyce

Cofactor induced dissociation of the multifunctional multisubunit EcoR124I investigated using electromobility shift assays, AFM and SPR

Popis výsledku anglicky

We have applied three techniques to the study of subunit assembly of the Type IC Restriction-Modification enzyme EcoR124I. This fully functional enzyme EcoR124I consists of a complex of the three subunits HsdR, HsdM and HsdS in a R2M2S1 stoichiometry, but is known to dissociate readily, releasing free HsdR and producing first an R-1-complex and then the core, DNA-binding methyltransferase (M2S1) complex. Analysis of the assembly pathway of this enzyme has previously employed gel retardation and Surface Plasmon Resonance (SPR), but the studies to date have not included the cofactors required for full enzyme activity. In this paper, we have also used atomic force microscopy (AFM)-based molecular volume measurements, and have analysed the effect of the cofactors ATP and AdoMet on enzyme stability and subunit assembly. We compare the data obtained from all three techniques and we show that they all give consistent results, but inherent differences in the methodologies provide additional information useful for the study of subunit assembly.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10606 - Microbiology

Projekt

<a href="/cs/project/GA204%2F07%2F0325" target="_blank" >GA204/07/0325: Identifikace aminokyselinových zbytků podjednotek HsdS a HsdR nutných pro správné sestavení restrikčně-modifikačního enzymu Typu I EcoR124I</a><br>

Návaznosti

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

Rok uplatnění

2017

Kód důvěrnosti údajů

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

Název periodika

RSC Advances

ISSN

2046-2069

e-ISSN

—

Svazek periodika

7

Číslo periodika v rámci svazku

61

Stát vydavatele periodika

GB - Spojené království Velké Británie a Severního Irska

Počet stran výsledku

10

Strana od-do

38737-38746

Kód UT WoS článku

000407442000072

EID výsledku v databázi Scopus

2-s2.0-85027237337