Добавил:
Upload Опубликованный материал нарушает ваши авторские права? Сообщите нам.
Вуз: Предмет: Файл:

Physics and chemistry of bioluminescence - Rajeev Ranjan, PhD

.pdf
Скачиваний:
26
Добавлен:
29.03.2016
Размер:
1.3 Mб
Скачать

Biological engineering Program

Physics and chemistry of bioluminescence

Rajeev Ranjan, PhD

Biological engineering Program

Compulsory course

Physics and chemistry of bioluminescence

(3 ECTS)

Diversity among luciferase class of enzyme, reaction mechanisms

Enzyme commission (EC) number

Different Luciferin structures, their role in luminescence emission

Introduction to Photoprotein(s)

Overview of bioluminescence: bioluminescence in nature

k

h

a b

c

e

g

i

l

 

 

 

 

d

m

f

j n

Introduction to luciferase: the enzyme that triggers the oxidation of the luciferin

Luciferase is a generic term

Belong to the class oxidoreductase and catalyse oxidation of respective substrates

Form product molecules in their electronically excited state which subsequently decay in form of light

Range of wavelength emission (visible) but generally in the range of 470-560 nm but is not limited in this range

Basic Bioluminescent systems

S.

Enzyme

Bioluminescent

Luciferase/MW(kDa)/

Substrate

No.

Commission

protein

Subunit

(Luciferin)

 

No.

 

 

 

 

1.

1.14.14.3

Bacterial luciferase

Bluc ≈ 79, dimer

Aliphatic

 

 

 

 

aldehyde

2.

1.13.12.7

Firefly luciferase

Fluc≈ 62, monomer

D-Luciferin

3.

1.13.12.5

Renilla luciferase

Rluc≈ 36, monomer

a

Coelenterazine

 

 

 

 

 

 

4.

1.13.12.18

Gonyaulax

LCF≈ 130, monomer

Open

 

 

luciferase

 

tetrapyrrole

5.

?

Fungi

?

b

ND

 

 

 

 

 

 

6.

1.13.12.6

Gaussia luciferase

Gluc≈ 20 monomer

a

Coelenterazine

 

 

 

 

 

 

7.

1.13.12.6

Metridia luciferase

Mluc≈ 24 monomer

a

Coelenterazine

 

 

 

 

 

 

8.

1.13.12.6

Vargula luciferase

Vhl≈ 62 monomer

a

Coelenterazine

 

 

 

 

 

 

9.

1.13.12.6

Cypridina luciferase

Cnl≈ 62 monomer

a

Coelenterazine

 

 

 

 

 

 

 

 

 

 

 

 

 

cPhotoprotein

 

 

1.

1.13.12.5

Aequorin

Aeq≈ 22 monomer

a

Coelenterazine

 

 

 

 

 

 

2.

1.13.12.6

Obelin

Obe≈ 22 monomer

a

Coelenterazine

 

 

 

 

 

 

aGroup of structurally related compounds, bNot determined, cLuciferase– luciferin complex

Bacterial luciferase (Bluc: EC 1.14.14.3)

Catalyzes oxidation of reduced flavin mononucleotide (FMNH2)

In the presence of luciferin (a long chain aldehyde) to yield the corresponding acid (a long chain acid), FMN, H2O and light (λmax: 490 nm)

EC 1. Oxidoreductase (A+ B → A + B)

EC 1.14. Acting on paired donors, with O2 as oxidant and incorporation or reduction of oxygen. The oxygen incorporated need not be derived from O2

EC 1.14.14. With reduced flavin or flavoprotein as one donor, and incorporation of one atom of oxygen into the other donor

EC 1.14.14.3 alkanal monooxygenase (FMN)

Bacterial luciferase (Bluc: EC 1.14.14.3)

The molecule of FMNH2 is deprotonated at N1 when bound to a luciferase molecule

Readily peroxidized at C4a to form Intermediate

Intermediate A reacts with a fatty aldehyde (such as dodecanal and tetradecanal) to form Intermediate

Intermediate B decomposes and yields the excited state of 4ahydroxyflavin (Intermediate C) and a fatty acid

Bacterial bioluminescence system (Bluc: EC 1.14.14.3)

Firefly luciferase (Fluc: EC 1.13.12.7)

Catalyzes oxidation of D-luciferin (Benzothiazole)

In the presence of Adenosine-5ꞌ-phosphate (ATP) to yield oxyluciferin (Oxidized benzothiazole), AMP, CO2 and light (λmax: 560 nm)

EC 1. Oxidoreductase (A+ B → A + B)

EC 1.13. Incorporation of oxygen into the substrate (oxygenases). The oxygen incorporated need not be derived from O2

EC 1.13.12. Incorporation of one atom of oxygen atom into the other donor

EC 1.13.12.7 Photinus-luciferin 4-monooxygenase (ATP-hydrolysing)

Firefly luciferase (Fluc: EC 1.13.12.7)

firefly luciferase (FLuc), beetle luciferin (LH2), oxyluciferin (OxyLH2), Mg-ATP, emitter oxyluciferin (OxyLH2), inhibitory side product dehydroluciferyl-AMP (L-AMP)

Two FLuc-catalyzed half reactions

Production of luciferyl-adenylate (LH2-AMP)

Reaction of LH2-AMP with O2 to produce OxyLH2 in an electronically excited state

Formation of a key dioxetanone intermediate

FLuc reaction is initiated when a presumed active site base

abstracts the C4 proton of LH2-AMP producing a carbanion

Тут вы можете оставить комментарий к выбранному абзацу или сообщить об ошибке.

Оставленные комментарии видны всем.

Соседние файлы в предмете Биофизика