Äîêóìåíò âçÿò èç êýøà ïîèñêîâîé ìàøèíû. Àäðåñ îðèãèíàëüíîãî äîêóìåíòà : http://lcc.chem.msu.ru/papers/biotech2007.pdf Äàòà èçìåíåíèÿ: Thu Nov 19 18:53:28 2009 Äàòà èíäåêñèðîâàíèÿ: Mon Oct 1 21:35:02 2012 Êîäèðîâêà:

Quantum Chemistry in Studies of Elementary Stages of Enzymatic Catalysis Alexander Nemukhin

Department of Chemistry M.V. Lomonosov Moscow State University Russian Federation

N.M. Emanuel Institute of Biochemical Physics Russian Academy of Sciences


The aim is to study mechanisms of chemical reactions in complex molecular environment by considering multidimensional potential energy surfaces in the QM/MM approach E = EQM + EMM (+ EQM/MM)

QM

MM-part

QM-part

MM


Software
QM MM QM: PC GAMESS (A. Granovsky) MM: TINKER (J. Ponder) Original QM/MM interfaces Current limitations: 100 atoms in QM 5000 atoms in MM

References on methodology and tests:
Nemukhin A., Grigorenko B., et al. // J. Phys. Chem. A, 2002, 106, 10663 Nemukhin A., Grigorenko B., Bochenkova A. et al. // Theochem, 2002, 581, 167 Nemukhin A., Grigorenko B., et al. // J. Comput. Chem., 2003, 24, 1410 Nemukhin A., Grigorenko B., et al. // J. Phys. Chem. B, 2003, 107, 2958 Nemukhin A., Grigorenko B., et al. // Phys. Chem. Chem. Phys., 2004, 6, 1031 Grigorenko B., Nemukhin A., et al. //J. Molec. Modeling, 2005, 11, 503 Grigorenko B., Rogov A., Nemukhin A. // J. Phys. Chem. B, 2006, 110, 4407 Rogacheva M., Bochenkova A., Nemukhin A., et al. // J. Phys. Chem. B, 2007, 111, 432


Hydrolysis of acetylcholine by acetylcholinesterase (AChE)
(Lushchekina S., Bochenkova A., Nemukhin A., Varfolomeev S. //to be submitted)

Serine hydrolase AChE rapidly hydrolyses the neurotransmitter

acetylcholine
H3C

O O N

CH3
+

to choline and acetic acid

CH3 CH3


Conclusions
Deacylation is a rate limiting stage
E, kcal/mol
TS 1

3.8

E, kcal/mol

8.4
TS 2 AE

ES

15.9
TI 2 TI 1

9.7 6.7
P

Acylation

Deacylation

One-proton transfer mechanism occurs at both stages: Glu does not abstract proton from His upon formation of tetrahedral intermediates

+
His

Glu


Serine proteases
(Nemukhin A., Grigorenko B., et al. //Theoret. Chem. Accounts, 2004, 111, 36)

Significance: ·Textbook example of enzymatic mechanism

·Design of novel inhibitors is a current task of biotechnology


ESINT1

INT1EA1

EA1EA2

ES

Complete catalytic cycle as a flow between stationary points on PES

EA2INT2 EP INT2EP


(Nemukhin A., Gariev I., Rogov A., Varfolomeev S. //Mendeleev Commun., 2006, 6, 290) The histogram of distribution of enzymes by the distances between the hydroxyl oxygen (O) from serine and nitrogen (N) from histidine: the PDB archive free enzymes enzymes with substrates/inhibitors
O
0.98 å

H

1.76 å

2.70 å Ser

N

His

2,2

2,4

2,6

2,8

3,0

3,2

3,4

3,6

3,8

4,0

R(O(Ser)-N(His)), å

The QM/MM calculations estimate the fraction of activation energy which could be gained upon compression of the enzyme due to substrate binding: ~ 3 kcal/mol or ~ 3 orders in the rate constant. Direct credentials to the Koshland's induced fit concept of enzymatic catalysis.


Sedolisin Catalysis
(Bravaya K., Bochenkova A., Grigorenko B., Nemukhin A., et al. //J. Chem. Theory & Comput., 2006, 2, 1168)

Significance: ·Design of enzymes active at unusual conditions (low pH, high T) ·Cleavage of insulin ·A novel enzymatic mechanism (confirmed by our simulations)

Ser287 Glu80 Asp84


The mechanism of the first stage ES EA shares features of that from serine proteases and aspartic proteases !

substrate substrate

ES

TI

EA

substrate


Enzymatic hydrolysis of guanosine triphosphate:
(Topol I., Nemukhin A., Grigorenko B., et al. //Biochim. Biophys. Acta, 2004, 1700, 125 Grigorenko B., Nemukhin A., et al. //Proteins, 2005, 60, 495 Grigorenko B., Shadrina M., Nemukhin A., et al. // Proteins, 2007, 66, 456)
GTP molecule
O C N C C N C N N N

GTP + H2O GDP + Pi
O P O O P OOOOO O P O C C O

C C

C C O O


Guanosine triphosphate (GTP) hydrolysis in Ras-GAP
Significance · One of the key biochemical reactions: provides a switch mechanism in controlling signal transduction including cell profileration and differentiation · Controversial viewpoints on the reaction mechanism

GTP


Computed energy profiles for water, Ras, Ras-GAP
25 20 15 10 E, kcal/mol 5 0 -5 -10 -15 -20 -25

20.0 16.0 7.0 4.3 -1.6 -2.0 14.0 10.5 4.6

-6.5 -9.2 -20.7

Reagents

TS1

Intermediate TS2

Products


ATP hydrolysis by myosin
(Grigorenko B., Nemukhin A., et al. //Proc. Natl. Acad. Sci. USA, accepted for publication)
ATP


Thanks to Bella Grigorenko (Moscow State University) Sergei Varfolomeev (Institute of Biochemical Physics) Sofia Lushchekina (Institute of Biochemical Physics) Alexander Moskovsky (Institute of Biochemical Physics) Anastasia Bochenkova (Moscow State University) Maria Shadrina (Moscow State University) Ksenia Bravaya (Moscow State University) Alexander Granovsky (Moscow State University) Alexander Rogov (Moscow State University) Igor Topol (National Cancer Institute at Frederick, USA) Stanley Burt (National Cancer Institute at Frederick, USA) Support from ·Russian Foundation for Basic Research ·Russian Academy of Sciences ·Advanced Biomedical Computing Center of the National Cancer Institute, USA