Бесплатный автореферат и диссертация по биологии на тему

Молекулярный и генетический анализ некоторых семейств гликозил-гидролаз микроорганизмов

ВАК РФ 03.00.03, Молекулярная биология

Содержание диссертации, кандидата биологических наук, Наумов, Даниил Геннадиевич

ВВЕДЕНИЕ

ОБЗОР ЛИТЕРАТУРЫ

Глава I. Гликозил-гидролазы: принципы классификации

1. Классификации, основанные на энзиматических свойствах гликозидаз

2. Классификация, основанная на структурном сходстве гликозидаз

Глава II. Системы утилизации сахарозы грамположительных бактерий

1. Система утилизации сахарозы Bacillus subtilis

1.1. Два пути утилизации сахарозы

1.2. Сахаразный и левансахаразный регулоны: сходство с ôgZ-опероном Escherichia coli

1.3. Леваназный оперон

1.4. Утилизация сахарозы у других штаммов Bacillus subtilis

2. Системы утилизации сахарозы у других представителей рода Bacillus

2.1. Bacillus stearothermophilus

2.2. Bacillus circulons

2.3. Paenibacillus (Bacillus) polymyxa

2.4. Bacillus licheniformis

2.5. Bacillus amyloliquefaciens

2.6. Bacillus halodurans

2.7. Другие виды Bacillus

2.8. Штаммы Bacillus неизвестной видовой принадлежности

3. Системы утилизации сахарозы у других немолочнокислых грамположительных бактерий

3.1. Staphylococcus

3.2. Clostridium 30 Ъ.Ъ. Bifidobacterium 31 3.4. Актиномицеты

4. Системы утилизации сахарозы молочнокислых бактерий

4.1. Streptococcus mutans

4.2. Другие представители рода Streptococcus

4.3. Lactococcus lactis

4.4. Leuconostoc mesenteroides

4.5. Lactobacillus и Pediococcus Ъ

Глава III. Гены ферментации мелибиозы Saccharomyces cerevisiae и других видов дрожжей

МАТЕРИАЛЫ И МЕТОДЫ

1. Бактериальные штаммы, среды и тестирование сбраживания сахарозы

2. Выделение ДНК из молочнокислых бактерий

3. Полимеразная цепная реакция

4. Методы генной инженерии

5. Среды для дрожжей и тестирование сбраживания мелибиозы

6. Штаммы дрожжей и методы генетической гибридизации

7. Молекулярное кариотипирование и Саузерн-гибридизация хромосом дрожжей

8. Компьютерный анализ аминокислотных последовательностей 57 РЕЗУЛЬТАТЫ И ОБСУЖДЕНИЕ

1. Скрининг коллекции штаммов с помощью ПЦР

2. Секвенирование фрагмента гена scrB Lactobacillus plantarum

3. Рестрикционное картирование локуса утилизации сахарозы

4. Сравнительный анализ последовательностей бактериальных сахараз

5. Генетическое картирование а-галактозидазных генов семейства MEL1-MEL11 в теломерах дрожжей Saccharomyces cerevisiae

6. Идентификация нового семейства а-галактозидазных генов MEL12-MEL14 у Saccharomyces cerevisiae

7. Генетическое картирование а-галактозидазных генов семейства MEL12-MEL14 в теломерах дрожжей Saccharomyces cerevisiae

8. Анализ аминокислотных последовательностей (З-фруктозидаз. Обнаружение (З-фруктозидазного суперсемейства

9. a-L-Арабиназы и P-D-ксилозидазы - гомологи p-фруктозидаз 110 10. Обсуждение результатов анализа структур белков фуранозидазного суперсемейства

ВЫВОДЫ

Заключение Диссертация по теме "Молекулярная биология", Наумов, Даниил Геннадиевич

выводы

1. У молочнокислых бактерий Lactobacillus plantarum NRRL В-4496 и Pediococcus pentosaceus NRRL В-11465 обнаружена сходная структура локуса утилизации сахарозы: дивергентно расположенные ген scrA, кодирующий сахарозо-специфичный второй фермент фосфотрансферазной системы, и гены scrB, scrR и agi, кодирующие ß-фруктозидазу, регуляторный белок и а-глюкозидазу соответственно. Сходство 5'-концевых частей генов scrB у L. plantarum и P. pentosaceus достигает 98.6%.

2. Проведено картирование в субтеломерных районах хромосом Saccharomyces cerevisiae девяти полимерных генов ферментации мелибиозы: ген MEL2 локализован в левом плече седьмой хромосомы (VII L), MEL3 - в XVI L, MEL4 - в XI L, MEL5 - в IV L, MEL6 - в XIIIR, MELI - в VI R, MEL8 - в XV R, MEL9 - в X R и MEL10 - в XIIR.

3. У штамма S. cerevisiae CBS 2888 обнаружено три гена ферментации мелибиозы MEL12-MEL14, представляющие новое дивергентное семейство а-галактозидазных генов. Проведено картирование этих генов в субтеломерных районах левых плечей хромосом S. cerevisiae: MELI2 в — IX, MEL13 - в XV и MEL14 - в X.

4. Обнаружена закономерность локализации полимерных генов семейств утилизации углеводов у S. cerevisiae. Гены ферментации сахарозы, мальтозы и мелибиозы, принадлежащие одному семейству (SUC, MAL, MELI-MELI 1 или MEL12-MEL14), никогда не находятся в противоположных концах одной и той же хромосомы (даже в разных штаммах).

5. Обнаружена гомология аминокислотных последовательностей ß-фруктозидаз, принадлежащих двум разным семействам гликозил-гидролаз: GH32 и GH68. Эти два семейства объединены в ß-фруктозидазное суперсемейство.

6. Обнаружена гомология белков ß-фруктозидазного суперсемейства с ß-D-ксилозидазами и a-L-арабинофуранозидазами семейств GH43 и GH62 гликозил-гидролаз. Оба семейства включены в состав ß-фруктозидазного суперсемейства. В состав этого суперсемейства также включена группа гипотетических белков, вероятно, кодирующих гликозидазы.

Библиография Диссертация по биологии, кандидата биологических наук, Наумов, Даниил Геннадиевич, Москва

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