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E. COLI. . ., . ., . .
(, )

E. coli . , pH . , . E. coli .
(EC 2.3.3.1, gltA) -- , - (AcCoA) (OAA) (Cit) (CoA) [1]: AcCoA + OAA = Cit + CoA.

. , E. coli in vitro. [2] , [3]. DBSolve7.0 [4]. . . E. coli: 1) - -- , [5] AcCoA [5]

. . . -- -- 2005, . 3, . 934 935 Mogilevskaya E. A. et. al. -- MCE -- 2005, vol. 3, p. 934 935

KCl. 0.1 KCl [5]. 0.1 M KCl E. coli, , KCl = 0. 2) : 2.24*106 [6]. 3) -- [7] [5]. , , , - [7], , [5]. 4) - -- , [8]. 5) -- , - [8]. 6) [8]. , pH [8]. 7) E. coli: Ac K m CoA = 0.11 mM;

K

= 0.7 mM (pH 8.0) [7]; K kcat = 4860 1/min (pH 8.0) [10]; pHopt = 7.3 [8].

OAA m AcCoA d

= 0.021 (pH 8.1, t = 21°C 0.1 KCl) [9]; (1)

1) . . , , AcCoA [8]. 935

8. , Part 8. Mathematical methods in biology, ecology and chemistry

, AcCoA . , , Ordered Bi Bi [11] AcCoA. . 1. - , -- AcCoA AcCoA OAA (. . 1), . , . , pH ( -), [2], , , (. 1). pH- (. . 6) , , .. pH- .

. 1. E. coli.

2) : (3) 1.
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. . . -- -- 2005, . 3, . 934 937 Mogilevskaya E. A. et. al. -- MCE -- 2005, vol. 3, p. 934 937

, . (3) 12 : 3 (1), 8 , in vitro . pH (kcat = 4860 1, pH 8.0 [10]) kcat 0 :

k

cat 0

KH H = k ca t * 1 + d 1 + H . H Kd 2

, 7. . AcCoA [7, 9]: (. 2,), AcCoA [9] (. 2), pH [8] (. 2). , (3) . . 2. :

KdH1 = 1e - 5 ; K K
AcCoA d

H d2

= 2e - 4 ; K = 0.58 .

AcCoA m

= 0.18 ; K

OAA m

= 0.04 ;

= 0.1; K

ATP i

-- - [7]:

K K

KG i1

= 0.015 ; K

KG i2

= 0.256 ;
NADH i2

NADH i1

= 3.3e - 4 ; K

= 8.4e - 3 .
937

8. , Part 8. Mathematical methods in biology, ecology and chemistry

. 2. pH, (3)

. 3.

. 3. , (3)

, . , 938

. . . -- -- 2005, . 3, . 934 939 Mogilevskaya E. A. et. al. -- MCE -- 2005, vol. 3, p. 934 939

[8]. , (3) (. 4).

. 4. . , (3)

E. coli. E. coli, (. 2). , E. coli -- , . pH , , pH. , [8], . , E. coli, . : 1. Neidhardt F.C. E. coli and Salm. typhimurium: Cellular and Molecular Biology. V.1. P.36.
939

8. , Part 8. Mathematical methods in biology, ecology and chemistry

2. - . . (. ..) --: - «», 1979. 282. 3. .. , .. , . , .. . Escherichia coli // . 2004. V.69. P.16251638. 4. Goryanin I., Hodgman T.C., and Selkov E. Mathematical simulation and analysis of cellular metabolism and regulation // Bioinformatics. 1999 V.15. P.749758. 5. D.S. Pereira, L.J. Donald, D.J. Hosfield, H.W. Duckworth. Active site mutants of Escherichia coli citrate synthase. Effects of mutations on catalytic and allosteric properties // J. Biol. Chem. 1994. V.269 N.1 P.412417. 6. Guynn R.W., Gelberg H.J., Veech R.L. Equilibrium Constants of the Malate Dehydrogenase, Citrate Synthase, Citrate Lyase, and Acetyl Coenzyme A Hydrolysis Reactions under Physiological Conditions // J. Biol. Chem. 1973. V.248. N.20. P.69576965. 7. Wright J.A., Sanwal B.D. Regulatory Mechanisms involving nicotinamide adenine nucleotides as allosteric effectors // J. Biol. Chem. 1971 V.246 N.6. P.16891699. 8. Jangaard N.O., Unkeless J., Atkinson D.E. The inhibition of Citrate Synthase by adenosine triphosphate // Biochim. Biophys. Acta. 1968. V.151. P.225235. 9. Faloona G.R., Srere P.A. Escherichia coli citrate synthase. Purification and the effect of potassium on some properties //Biochemistry. 1969 V.8. N.11. P.44974503. 10. Donald L.J., Crane B.R., Andersone D.H., Duckworth H.W. The role of cysteine 206 in allosteric inhibition of Escherichia coli citrate synthase // J. Biol. Chem. 1991. V.266. N.31. P.2070920713. 11. Cleland W.W. The kinetics of enzyme-catalyzed reactions with two or more substrates or products. I. Nomenclature and rate equations // Biochim. Biophys. Acta. 1963. V. 67, P.104137. 12. Peng L., Shimizu K. Global metabolic regulation analysis for Escherichia coli K12 based on protein expression by 2dimensional electrophoresis and enzyme activity measurement // Appl. Microbiol. Biotechnol. 2003 V.61. P.163178. 13. Padan E, Zilberstein D, Schuldiner S. pH homeostasis in bacteria // Biochim. Biophys. Acta. 1981 V.650. P.151166.
940

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1. .
1) : [6], . (. 1) , k3. , , . , -- , . , : K H H ATP V = CS * k1k2k3 * AcCoA*OAA ( k-1k3 + k-1k-2 + k2k3 *OAA) 1+ d1 + H *1+ ATP + HK Ki d2

+ AcCoA * ( k1k3 + k1k

+ H Kd1 H KG NADH + H + KG + NADH , + k1k2 * AcCoA * OAA * 1 + H K d 2 Ki 2 Ki 2
-2

)

KH H KG NADH * 1 + d 1 + H + KG + NADH H K d 2 Ki1 Ki1

(1)

k1, 1, k2, 2, k3 -- , CS -- , K dH1 , K dH2 , K iATP , K iKG , K iKG , K iNADH , K iNADH -- 1 2 1 2 . . 1 . 2) , ( , , .) , (1) ,
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8. , Part 8. Mathematical methods in biology, ecology and chemistry

(1):
Vmax = CS * k
3

K dH1 H +H 1 + H Kd 2

;

K

AcCoA m

=

k3 ;K k1

OAA m

=

k3 + k-2 . k2

(2)

3) : (2), , : K H H ATP O Ac V = CS * kcat 0 * AcCoA*OAA ( KdAcCoA * KmAA + Km CoA *OAA) 1+ d1 + H *1+ ATP + HK Ki d2
KH H KG NADH + AcCoA * K OAA * 1 + d1 + + + m H KG K NADH H K K d2 i1 i1 +

KH H KG NADH + AcCoA * OAA * 1 + d 1 + H + KG + NADH , H K d 2 Ki 2 Ki 2

(3)

k

cat 0

= k3 ; K

AcCoA d

=

k-1 . k1

2. E. coli.
(SA) E. coli [12]: SA = 0.25 /*

.

, 1 E. coli 5.5 [1] : Vmax = 45.5 /. , . , pH 7.3 [13], pH- :

942

. . . -- -- 2005, . 3, . 934 943 Mogilevskaya E. A. et. al. -- MCE -- 2005, vol. 3, p. 934 943 k
pH7.3 cat

=k

cat 0

, E. coli, :
CS
acetate acetat = Vmax e / k pH 7.3 cat

K dH1 H 1e - 5 1e - 4.3 -1 + H = 9941 1 + + 1 + = 6966 . H Kd 2 1e - 4.3 2.2e - 4

= 6.5 .

, E. coli, (SA = 0.05 /* [12], Vmax = 9 /), , : glucos pH CSglucose = Vmax e / kcat 7.3 = 1.3 .

KINETIC MODEL OF E. COLI CITRATE SYNTHASE Mogilevskaya E. A., Lebedeva G. V., Demin O. V.
(Russia, Moscow)

Kinetic model of E. coli Citrate Synthase has been constructed on the base of literature experimental data. Rate equation has been derived. Inhibitors and pH influence on the enzyme functionning was taken into account. All unknown parameters of rate equation have been determined. Citrate Synthase concentrations in E. coli cells grown aerobically on acetate and glucose have been defined using data on cell extracts specific activities.

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