Transferazele

Aminotransferazele

Aspartat-aminotransferaza
(L-aspartat: -cetoglutarat- aminotransferaza

alanin-aminotransferazei
(L-alanin--cetoglutarat- aminotransferaza
glutamat-piruvat transaminaza).

Trascetolaza (sedoheptulozo-7-fosfat:Dgliceraldehid-3-fosfat-glicol-aldehid-transferaza)

ATP fosfohidrolaza

Timidat sintaza (5,10-metilentetrahidrofolat:dUDP-metiltransferaza)

Hidrolazele-Fosfolipazele

Amilazele

Izomerazele
Catalizeaza rearanjarea unor legaturi dintr-o molecula

Racemaze
Izomerazele

Epimeraze
Cis-Trans Izomeraze
Tautomeraze

Racemazele
Amino-acid racemazele
L-aminoacid

D-aminoacid
Alanil-racemaza

Racemaze

Metionin-racemaza
Glutamat-racemaza

Alanil-racemaza

Piridoxal
fosfat
Bacillus anthracis

Mandelat-racemaza

Cis-trans izomeraze

Enoil-CoA
izomeraza

Prolil cis-trans izomerazele

Fanghnel J, Fischer G (2004)

Front Biosci 9:3453

Coenzima B12

Protein disulfide izomeraza

>10 proteine de tip PDI in tesuturile umane

ox

red
Gruber, CW et al. TIBS, 2006, 31: 455-464.

Sintetaze (ligaze)

Aminoacil-adenilat
(anhidrida mixta)

fiecare molecula ARNt

un aminoacid

Aminoacil-ARNt

Acetil-CoA sintetaza

L-aspartat:amoniac ligaza

Acetil-CoA carboxilaza

Carbamilfosfat sintetaza I

Carbamilfosfat sintetaza II

Arginino-succinat sintetaza

Studiul enzimelor -obiective:

1) determinarea masei moleculare relative
(KDa, 1 Dalton = 1/12 din masa atomica
12C =1,66 x 10-24 g);
2) determinarea compozitiei aminoacizilor;
3) determinarea structurii primare;
4) determinarea structurii secundare si
tertiare;

Determinarea masei
moleculare
a proteinelor

Mr =

RTs
D (1-v)

Mr =

2RT
(1-v) 2

dlnc
dr2

Gel filtrarea
Kd =

Ve - Vo
Vi - Vo

MW/kDa

1000

100

10
0

20

40

60

80

Volume/ml

100

120

Ex
co tra
nt ct
in c
e ar
G en e
FP zi
m
p
G ur a
FP a
St den
an
a
da tur
at
rd
a

SDS PAGE

1000

66 kDa
45 kDa

M W/kDa

97 kDa

100

30 kDa
10
0

20 kDa
14 kDa

10

20
distance/mm

30

Determinarea Mr a
proteinelor si peptidelor
Tris-glycine SDS PAGE (Laemmli)
7.5% 40-400 kDa
12.5% 20-100 kDa
20% 3-25 kDa
Tris-Tricine SDS PAGE (Schaegger)
16.5% 2-25 kDa

Structura insulinei
(hormon)

PDB 3BTK

Asp102
His57
Ser195

Resturile catalitice din tripsina

set

feature

inhibitor

examples

function

Serine protease

active site serine

H57, D102, S195

fluorophosphates

trypsin
thrombin
plasmin
coccoonase
subtilisin
acrosin

digestion
blood coagulation
lysis of blood clots
mechanical
digestion
sperm penetration

Cysteine protease

active site cysteine

C25, H159, N175

iodoacetate

papain
strept. proteinase
cathepsin B

digestion
digestion
intracell. digestion

Acid protease

acidic pH optimum
D32, D215

diazoketones

pepsin
chymosin

digestion
milk coagulation

Metalloproteases

Zn2+, E270
Zn2+, Ca2+
E143, H231

o-phenanthroline
o-phenanthroline

carboxypeptidase
thermolysin

digestion
digestion