Variations in the enzymatic activity of S1-type nucleases results from differences in their active site structures

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F60077344%3A_____%2F23%3A00584258" target="_blank" >RIV/60077344:_____/23:00584258 - isvavai.cz</a>

Výsledek na webu

<a href="https://www.sciencedirect.com/science/article/pii/S0304416523001228?via%3Dihub" target="_blank" >https://www.sciencedirect.com/science/article/pii/S0304416523001228?via%3Dihub</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1016/j.bbagen.2023.130424" target="_blank" >10.1016/j.bbagen.2023.130424</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Variations in the enzymatic activity of S1-type nucleases results from differences in their active site structures

Popis výsledku v původním jazyce

Background: S1-like nucleases are widespread enzymes commonly used in biotechnology and molecular biology. Although it is commonly believed that they are mainly Zn2+ dependent acidic enzymes, we have found that numerous members of this family deviate from this rule. Therefore, in this work, we decided to check how broad is the range of non-zinc-dependent S1-like nucleases and what is the molecular basis of their activities. Methods: S1-like nucleases chosen for analysis were achieved through heterologous expression in appropriate eukaryotic hosts. To characterize nucleases active-site properties, point mutations were introduced in selected positions. The enzymatic activities of wild-type and mutant nucleases were tested by in-gel nuclease activity assay. Results: We discovered that S1-like nucleases encoded by non-vascular plants and single-celled protozoa, like their higher plant homologues, exhibit a large variety of catalytic properties. We have shown that these individual properties are determined by specific non-conserved active site residues. Conclusions: Our findings demonstrate that mutations that occur during evolution can significantly alter the catalytic properties of S1-like nucleases. As a result, different ions can compete for particular S1-type nucleases active sites. This phenomenon undermines the existing classification of S1-like nucleases. General significance: Our findings have numerous implications for applications and understanding the S1-like nucleases biological functions. For example, new biotechnological applications should take into account their unexpected catalytic properties. Moreover, these results demonstrate that the trinuclear zinc-based model commonly used to characterize the catalytic activities of S1-like nucleases is insufficient to explain the actions of non-zinc-dependent members of this family.

Název v anglickém jazyce

Variations in the enzymatic activity of S1-type nucleases results from differences in their active site structures

Popis výsledku anglicky

Background: S1-like nucleases are widespread enzymes commonly used in biotechnology and molecular biology. Although it is commonly believed that they are mainly Zn2+ dependent acidic enzymes, we have found that numerous members of this family deviate from this rule. Therefore, in this work, we decided to check how broad is the range of non-zinc-dependent S1-like nucleases and what is the molecular basis of their activities. Methods: S1-like nucleases chosen for analysis were achieved through heterologous expression in appropriate eukaryotic hosts. To characterize nucleases active-site properties, point mutations were introduced in selected positions. The enzymatic activities of wild-type and mutant nucleases were tested by in-gel nuclease activity assay. Results: We discovered that S1-like nucleases encoded by non-vascular plants and single-celled protozoa, like their higher plant homologues, exhibit a large variety of catalytic properties. We have shown that these individual properties are determined by specific non-conserved active site residues. Conclusions: Our findings demonstrate that mutations that occur during evolution can significantly alter the catalytic properties of S1-like nucleases. As a result, different ions can compete for particular S1-type nucleases active sites. This phenomenon undermines the existing classification of S1-like nucleases. General significance: Our findings have numerous implications for applications and understanding the S1-like nucleases biological functions. For example, new biotechnological applications should take into account their unexpected catalytic properties. Moreover, these results demonstrate that the trinuclear zinc-based model commonly used to characterize the catalytic activities of S1-like nucleases is insufficient to explain the actions of non-zinc-dependent members of this family.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10608 - Biochemistry and molecular biology

Projekt

—

Návaznosti

I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Rok uplatnění

2023

Kód důvěrnosti údajů

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

Název periodika

Biochimica et Biophysica Acta-General Subjects

ISSN

0304-4165

e-ISSN

1872-8006

Svazek periodika

1867

Číslo periodika v rámci svazku

10

Stát vydavatele periodika

NL - Nizozemsko

Počet stran výsledku

9

Strana od-do

130424

Kód UT WoS článku

001058918800001

EID výsledku v databázi Scopus

2-s2.0-85165714520