Russian Journal of Biological Physics and Chemisrty Russian Journal of Biological Physics and Chemisrty АКТУАЛЬНЫЕ ВОПРОСЫ БИОЛОГИЧЕСКОЙ ФИЗИКИ И ХИМИИ 2499-9962 54262 МЕДИЦИНСКАЯ БИОФИЗИКА И БИОФИЗИЧЕСКАЯ ХИМИЯ MEDICAL BIOPHYSICS AND BIOPHYSICAL CHEMISTRY МЕДИЦИНСКАЯ БИОФИЗИКА И БИОФИЗИЧЕСКАЯ ХИМИЯ INVESTIGATIONS OF ACCESSIBILITY OF T2/T3 COPPER CENTRE OF TWO-DOMAIN LACCASE FROM STREPTOMYCES GRISEOFLAVUS AC-993 ИССЛЕДОВАНИЕ ДОСТУПНОСТИ Т2/Т3 ЦЕНТРА ДВУХДОМЕННОЙ ЛАККАЗЫ ИЗ STREPTOMYCES GRISEOFLAVUS AC-993 Тищенко С В Tishchenko S V sveta@vega.protres.ru Костарева О С Kostareva O S Михайлина А М Mikhaylina A O Лисов А А Lisov A V Коляденко И А Kolyadenko I A Габдулхаков А Г Gabdulkhakov A G Институт белка РАН ru Institute of Protein Research, Russian Academy of Sciences ru Институт белка РАН ru Institute of Protein Research, Russian Academy of Sciences ru Институт белка РАН ru Institute of Protein Research, Russian Academy of Sciences ru Институт биохимии и физиологии микроорганизмов РАН ru Skryabin Institute of Biochemistry and Physiology of Microorganisms, Russian Academy of Sciences ru Институт белка РАН ru Institute of Protein Research, Russian Academy of Sciences ru Институт белка РАН ru Institute of Protein Research, Russian Academy of Sciences ru 25 03 2018 25 03 2018 3 1 163 167 20 03 2018 20 03 2018 https://rusjbpc.ru/en/nauka/article/54262/view

Лакказы (EC 1.10.3.2) - широко распространённые в природе «голубые» медьсодержащие оксидоредуктазы, являются перспективным объектом для биотехнолологической индустрии. Субстратами этих ферментов являются различные фенольные соединения, лакказы обнаружены в высших растениях, некоторых насекомых, грибах и бактериях. Лакказы содержат четыре атома меди, организованные в 3 сайта: Т1, Т2 и Т3. В бактериях, наряду с трёхдоменными лакказами, имеются двухдоменные лакказы (2DLac), которые имеют ряд функциональных преимуществ. 2DLac активны в нейтральных и щелочных значениях рН, обладают повышенной термостабильностью и устойчивы к действию различных ингибиторов. Каталитический механизм двухдоменных лакказ интенсивно изучается, однако, принципы субстрат/продуктного транспорта до сих пор не вполне понятны. В данной работе представлен сравнительный анализ каталитической активности и кристаллических структур рекомбинантной 2DLac из Streptomyces griseoflavus Ac-993 и её мутантных форм с заменами His165 на Phe и Ala. Консервативный для 2DLac His165 принадлежит второй координационной сфере и располагается близко к поверхности белка. Мы предполагаем, что движение имидазольного кольца His165 может «открывать» или «закрывать» один из субстрат-продуктных каналов, ведущих к Т2/Т3 центру лакказы S. griseoflavus .

Laccases (EC 1.10.3.2) are "blue" copper-containing oxidoreductases widely distributed and highly important for biotechnology. The substrates of these enzymes are various phenolic compounds. Laccases are found in higher plants, some insects, fungi and bacteria. Laccases contain four copper atoms organized in 3 sites: Т1, Т2 и Т3. In bacteria, along with three-domain laccases, there are two-domain laccases (2DLac), which have a number of functional advantages. 2DLac are active at neutral and alkaline pH values, have increased thermostability and are resistant to the action of various inhibitors. The catalytic mechanism of the two-domain laccase is intensively studied; however, the principles of substrate/product transport are still not fully understood. This paper presents a comparative analysis of the catalytic activity and crystal structures of recombinant 2DLac from Streptomyces griseoflavus Ac-993 and its mutant forms with His165 substitutions on Phe and Ala. His165 is conservative for 2Dlac, it belongs to the second coordination sphere and is located near the surface of the protein. We assume that the movement of the imidazole ring His165 can "open" or "close" one of the substrate-product channels leading to the T2 /T3 center of laccase from S. griseoflavus.

двухдоменные лакказы Streptomyces griseoflavus кристаллические структуры Т2/Т3 центр субстратные каналы two-domain laccases Streptomyces griseoflavus crystal structures T2 / T3 center substrates channels

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