Russian Journal of Biological Physics and Chemisrty Russian Journal of Biological Physics and Chemisrty АКТУАЛЬНЫЕ ВОПРОСЫ БИОЛОГИЧЕСКОЙ ФИЗИКИ И ХИМИИ 2499-9962 54579 МЕДИЦИНСКАЯ БИОФИЗИКА И БИОФИЗИЧЕСКАЯ ХИМИЯ MEDICAL BIOPHYSICS AND BIOPHYSICAL CHEMISTRY МЕДИЦИНСКАЯ БИОФИЗИКА И БИОФИЗИЧЕСКАЯ ХИМИЯ Modeling mutant human inosine triphosphate pyrophosphohydrolase P32T-ITPA and potential regulatory chemical modifications of the enzyme Моделирование мутантной формы инозин трифорсфат пирофосфогидролазы человека Р32Т-ITPA и потенциальные регуляторные химические модификации фермента Колтовая Н А Koltovaya N A koltovaya@jinr.ru Объединенный институт ядерных исследований ru Joint Institute for Nuclear Research ru 25 12 2020 25 12 2020 5 4 637 643 20 12 2020 20 12 2020 https://rusjbpc.ru/en/nauka/article/54579/view

Аналоги оснований относятся к мощным антиметаболитам и опасным мутагенам, образующимся эндогенно в условиях окислительного стресса, воспаления и аберрантного синтеза нуклеотидов. Инозин трифосфатпирофосфогидролаза человека (ITPA) гидролизует трифосфаты неканонических пуриновых оснований (ИТФ, дИТФ, XТФ, дХТФ) и, таким образом, регулирует пул нуклеотидов и защищает клетки от повреждений ДНК. Полиморфная аллель с.94C>A, характеризующаяся заменой Р32Т, приводит к утрате ферментативной активности в эритроцитах и накоплению ИТФ. Мутантная аллель P32T-ITPA ассоциирует с неблагоприятной реакцией на пуриновые аналоги, используемые в качестве препаратов при трансплантации органов, лечении рака крови и воспалительных заболеваний кишечника. Механизм инактивации мутантной формы P32T-ITPA не известен. Моделирование мутантной формы фосфогидролазы показало, что происходит ослабление связи между двумя субъединицами [1]. Действительно, согласно экспериментальным данным мутантный фермент менее стабилен, но активен in vitro [2]. Выдвинутая в литературе гипотеза о выталкивании гидрофобного остатка наружу, что служит сигналом для деградации белка [3], при моделировании мутантного фермента не подтвердилась [1]. Для понимания механизма инактивации in vivo необходимо более детальное рассмотрение молекулярных механизмов экспрессии гена. В процессе транскрипции гена происходит альтернативный сплайсинг и образование трех вариантов, причем в разных тканях пропорции этих вариантов разнятся. При белковом электрофорезе наблюдаются множественные полосы [4]. В данной работе, используя программы, определяющие потенциальные сайты химических модификаций, удалось определить сайты фосфорилирования, убиквитинирования и сумоилирования. Предполагается, что химические модификации участвуют в регуляции локализации белка и его активности. Полученные результаты позволяют планировать дальнейшую экспериментальную проверку наличия модифицированных форм и моделирование влияния химических модификаций на активность фермента.

Base analogs are powerful endogenous antimetabolites and dangerous mutagens forming under oxidative stress, inflammation and aberrant nucleotide synthesis. Human inosine triphosphate pyrophosphohydrolase (ITPA) catalyzes the hydrolysis of triphosphates of non-canonic purine bases (ITP, dITP, XTP, dXTP) and thus regulates the nucleotide pool and protects cells from DNA damage. The polymorphic allele c.94C>A, which encodes the enzyme with P32T substitution, causes enzymatic activity loss in erythrocytes and ITP accumulation. An association is observed between the mutant allele P32T-ITPA and adverse sensitivity to purine base analog drugs that are used in the treatment of acute lymphoblastic leukemia and inflammatory intestinal diseases and for preventing organ rejection after transplantation. The inactivation mechanism of the polymorph P32T-ITPA enzyme is unknown. A simulation of the enzyme's mutant form showed weakening of the interconnection between the two subunits [1]. Indeed, the experimental data indicate that the mutant enzyme is less stable but active in vivo [2]. The assumption made in the literature that the hydrophobic residue is pushed outside, which is a signal for proteasome degradation, has not been confirmed in silico [1]. To understand the mechanism of inactivation in vivo, it is necessary to examine the molecular mechanisms of gene expression in more detail. It has been found that alternative splicing and formation of three forms is possible; the latter's proportion being tissue specific. Numerous strips are observed in protein electrophoresis [4]. In the present work, using special programs that identify the potential chemical modification sites, we have determined the potential sites of phosphorylation, ubiquitination, and SUMOliation. It is assumed that chemical modifications participate in the regulation of the enzyme's location and activity. Hypothetic mechanisms are suggested. The results allow us to carry out a further experimental check of the existence of modified forms and to simulate the impact of chemical modifications on enzyme activity.

 однонуклеотидный полиморфизм гена ITPA аллельная форма инозин трифосфатпирофосфогидролаза Р32Т-ITPA компьютерное моделирование потенциальные сайты химических модификаций single nucleotide polymorphism ITPA gene mutant inosine triphosphatase P32T-ITPA computer simulation potential sites of regulatory chemical modifications

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