Substrate Specificity Diversity of Human Terminal Deoxynucleotidyltransferase May Be a Naturally Programmed Feature Facilitating Its Biological Function
Научная публикация
Журнал |
International Journal of Molecular Sciences
, E-ISSN: 1422-0067
|
Вых. Данные |
Год: 2024,
Том: 25,
Номер: 2,
Номер статьи
: 879,
Страниц
:
DOI:
10.3390/ijms25020879
|
Ключевые слова |
: human terminal deoxynucleotidyltransferase; V(D)J recombination; substrate specificity;
dNTP recognition; metal ion; cofactor; enzyme kinetics |
Авторы |
Kuznetsova Aleksandra A.
1
,
Senchurova Svetlana I.
1
,
Gavrilova Anastasia A.
1
,
Tyugashev Timofey E.
1
,
Mikushina Elena S.
1
,
Kuznetsov Nikita A.
2,1
|
Организации |
1 |
Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of Russian Academy of Sciences (SB RAS), 8 Prospekt Akad. Lavrentyeva, Novosibirsk 630090, Russia
|
2 |
Department of Natural Sciences, Novosibirsk State University, 2 Pirogova Str., Novosibirsk 630090, Russia
|
|
Информация о финансировании (2)
1
|
МИНИСТЕРСТВО НАУКИ И ВЫСШЕГО ОБРАЗОВАНИЯ РОССИЙСКОЙ ФЕДЕРАЦИИ
|
ПФНИ РФ (2021-2030) 0245-2021-0009
|
2
|
Российский научный фонд
|
РНФ №21-64-00017
|
Terminal 20
-deoxynucleotidyl transferase (TdT) is a unique enzyme capable of catalysing
template-independent elongation of DNA 30
ends during V(D)J recombination. The mechanism
controlling the enzyme’s substrate specificity, which is necessary for its biological function, remains
unknown. Accordingly, in this work, kinetic and mutational analyses of human TdT were performed
and allowed to determine quantitative characteristics of individual stages of the enzyme–substrate
interaction, which overall may ensure the enzyme’s operation either in the distributive or processive
mode of primer extension. It was found that conformational dynamics of TdT play an important
role in the formation of the catalytic complex. Meanwhile, the nature of the nitrogenous base
significantly affected both the dNTP-binding and catalytic-reaction efficiency. The results indicated
that neutralisation of the charge and an increase in the internal volume of the active site caused a
substantial increase in the activity of the enzyme and induced a transition to the processive mode in
the presence of Mg2+ ions. Surrogate metal ions Co2+ or Mn2+ also may regulate the switching of
the enzymatic process to the processive mode. Thus, the totality of individual factors affecting the
activity of TdT ensures effective execution of its biological function.