Parameters analysis and correlation coefficient between EFMs

Chapter 4 Model verification and validation

4.2.1 Parameters analysis and correlation coefficient between EFMs

The parameters have a primary role in mathematical model and greatly influence how the cell behavior be simulated.

About the control effect of parameters, Figure 4.1 illustrate the strange of parameters can be separated to a subset that has high coefficient, although the parameters didn’t have high coefficient for every metabolite, tend to prefer high coefficient. Because of the trend, select one of mode that correspond high coefficient parameter’s shown in Figure 4.4.Compare with Figure 2.11 and Figure 2.12, the reactions are a lot of different such as produce acetate and succinate ,and no ethanol produce. Studies should be undertaken to determine the frequency of each reactions operating shown in Table 4.1.the high frequency reactions such as F16BP = F6P, GL6P = 6PG, 6PG + NADP = R5P + CO2 + NADPH, and glycolysis pathway are more consistent than the fermentative pathway , that explain the major trunk pathway and branch pathway in high influence EFMs.

Table 4.1 Reaction frequency of EFMs for glycerol fermentation plus tryptone: 33~133

GG1 GG3 GG4 GG11 GG12 GG13 PPP2 PPP3 TCA1 TCA2r FR1

0 0 1 0.87 0 0.48 1 1 0.95 0.95 0.58

FR2 FR3 ANA1 ANA2 ANA3 FEM1 FEM2 FEM7 FEM8 FEM3 FEM5

0.63 0.63 1 0.19 0 0.49 0.19 0.25 0.25 0.13 0.12

FEM6 FEM4 FEM9 EDP1 EDP2 XYL1 XYL2 GAL1 MAN1 MAN2 ARA1

0.25 0.49 0.13 0 0 0 0 0 0 0 0

BIO OPM4r FC2 TRA1 TRA2 TRA3 TRA4 TRA5 TRA6 TRA7 GLB1

0.95 0.51 0 0.25 0.44 0.95 0.13 0.69 0 0.55 0.24

GLB2 TRA8 TRA9 TRA10 GLYD1 GLYD2 GLYD3 GLYD4 OPM3 TRYP GG2r

0.24 0 0 0 1 0 1 1 0 1 1

GG5r GG6r GG7r GG8r GG9r GG10r PPP1 PPP4r PPP5r PPP6r PPP7r

1 1 1 1 1 1 1 0.69 1 0.68 0.68

PPP8r TCA3r TCA4 FC1r

0.68 0.95 0.95 1

Moreover, the reaction rate is equal to parameters multiply elementary flux mode, the parameter value is an important characteristic of reaction rate. Figure 4.2 depict the parameters value express in logarithm and normal bar chart. It alteration of parameters in low coefficient part is small compare with high coefficient part in logarithm bar chart, and the average value of parameters in low coefficient part is larger than high coefficient part.

0.00E+00

Figure 4.1 Coefficient of parameters and metabolites concentration: glycerol fermentation plus tryptone (A) tryptone (B) Ethanol (C) Succinate (D) Glycerol (E) NH3 (F) Formate (G) Biomass (H) Lactate (I) CO2.

Furthermore, we investigate the correlation coefficient of each EFMs, that can realize that every EFMs are similar or not. Figure 4.3 shows the correlation coefficient in glycerol fermentation plus tryptone, the red point represent high correlation coefficient, yellow point represent low correlation coefficient, and green represent negative correlation coefficient.

There are many red square that reveal this part of EFMs is similar shown in Figure 4.3, and we collect three parts of EFMs that have high correlation coefficient.

(A)

Figure 4.2 Parameters distribution : glycerol fermentation plus tryptone .

EFM1 EFM2 EFM3 EFM4 EFM5 EFM6 EFM7 EFM8 EFM9 EFM10 EFM11 EFM12 EFM13 EFM14 EFM15 EFM16 EFM17

. . . .

Each of the three parts EFMs that we select one to compare with early work shown in Figure 4.5, Figure 4.6, and Figure 4.7. In Figure 4.5, the major product is acetate. In Figure 4.6, the major products are ethanol, biomass, and succinate. In Figure 4.7, it have the most less reaction that only convert glycerol to succinate and ethanol ,but this EFMs didn’t growing biomass. Above all, the cluster of EFMs matrix provides a possible thinking that performs the a few EFMs to stand for whole systems EFMs.

The frequency of high correlation coefficient part shown in Table 4.2-4.4, focus on mode 151 – mode 282, there is none of the reaction about biomass growing, and the less flow in pentose phosphate pathway and ratio of ethanol in fermentative produce is increased. That cluster of EFMs stand for high yields EFMs lead us to further research on the question of how to balance the cell growing and increase the ethanol yields.

Table 4.2 Reaction frequency of EFMs for glycerol fermentation plus tryptone: 1~32

GG1 GG3 GG4 GG11 GG12 GG13 PPP2 PPP3 TCA1 TCA2r FR1

0 0 1 0.87 0 0.47 0 0 1 1 0.57

FR2 FR3 ANA1 ANA2 ANA3 FEM1 FEM2 FEM7 FEM8 FEM3 FEM5

0.63 0.63 1 0.2 0 0.53 0.17 0.23 0.23 0.13 0.13

FEM6 FEM4 FEM9 EDP1 EDP2 XYL1 XYL2 GAL1 MAN1 MAN2 ARA1

0.27 0.53 0.13 0 0 0 0 0 0 0 0

BIO OPM4r FC2 TRA1 TRA2 TRA3 TRA4 TRA5 TRA6 TRA7 GLB1

1 0.53 0 0.27 0.4 1 0.13 0.7 0 0.53 0.27

GLB2 TRA8 TRA9 TRA10 GLYD1 GLYD2 GLYD3 GLYD4 OPM3 TRYP GG2r

0.27 0 0 0 1 0 1 1 0 1 1

GG5r GG6r GG7r GG8r GG9r GG10r PPP1 PPP4r PPP5r PPP6r PPP7r

1 1 1 1 1 1 0 1 1 1 1

PPP8r TCA3r TCA4 FC1r

1 1 1 1

Table 4.3 Reaction frequency of EFMs for glycerol fermentation plus tryptone: 33~133

GG1 GG3 GG4 GG11 GG12 GG13 PPP2 PPP3 TCA1 TCA2r FR1

0 0 1 0.87 0 0.48 1 1 0.95 0.95 0.58

FR2 FR3 ANA1 ANA2 ANA3 FEM1 FEM2 FEM7 FEM8 FEM3 FEM5

0.63 0.63 1 0.19 0 0.49 0.19 0.25 0.25 0.13 0.12

FEM6 FEM4 FEM9 EDP1 EDP2 XYL1 XYL2 GAL1 MAN1 MAN2 ARA1

0.25 0.49 0.13 0 0 0 0 0 0 0 0

BIO OPM4r FC2 TRA1 TRA2 TRA3 TRA4 TRA5 TRA6 TRA7 GLB1

0.95 0.51 0 0.25 0.44 0.95 0.13 0.69 0 0.55 0.24

GLB2 TRA8 TRA9 TRA10 GLYD1 GLYD2 GLYD3 GLYD4 OPM3 TRYP GG2r

0.24 0 0 0 1 0 1 1 0 1 1

GG5r GG6r GG7r GG8r GG9r GG10r PPP1 PPP4r PPP5r PPP6r PPP7r

1 1 1 1 1 1 1 0.69 1 0.68 0.68

PPP8r TCA3r TCA4 FC1r

0.68 0.95 0.95 1

Table 4.4 Reaction frequency of EFMs for glycerol fermentation plus tryptone: 151~282

GG1 GG3 GG4 GG11 GG12 GG13 PPP2 PPP3 TCA1 TCA2r FR1

0 0.19 0.34 0.76 0.19 0.4 0.15 0 0.35 0.35 0.53

FR2 FR3 ANA1 ANA2 ANA3 FEM1 FEM2 FEM7 FEM8 FEM3 FEM5

0.57 0.57 0.81 0.3 0.19 0.41 0.13 0.28 0.28 0.05 0.19

FEM6 FEM4 FEM9 EDP1 EDP2 XYL1 XYL2 GAL1 MAN1 MAN2 ARA1

0.28 0.41 0.09 0.15 0.15 0 0 0 0 0 0

BIO OPM4r FC2 TRA1 TRA2 TRA3 TRA4 TRA5 TRA6 TRA7 GLB1

0 0.45 0.19 0.28 0.4 0 0.05 0.67 0 0.54 0.35

GLB2 TRA8 TRA9 TRA10 GLYD1 GLYD2 GLYD3 GLYD4 OPM3 TRYP GG2r

0.35 0 0 0 1 0 1 1 0.19 0.91 0.15

GG5r GG6r GG7r GG8r GG9r GG10r PPP1 PPP4r PPP5r PPP6r PPP7r

0.15 0.98 0.98 0.98 0.98 0.98 0.15 0 0 0 0

PPP8r TCA3r TCA4 FC1r

0 0.35 0 0.15

Figure 4.4 The mode No.81 in glycerol fermentation in medium plus tryptone.

Figure 4.5 The mode No.18 in glycerol fermentation in medium plus tryptone.

Figure 4.6 The mode No.100 in glycerol fermentation in medium plus tryptone.

In other condition, glycerol fermentation plus furmarate, there are also high coefficient subset and low high coefficient subset parameters with metabolites concentration.. Although the high coefficient parameters modes influence averagely, it have less effect to the exception metabolite. Using these variations, the extreme condition and phenomenon can be revealed.

So the cluster of similar EFMs can not use few amounts of EFMs to represent the whole systems.

1 115 229 343 457 571 685 799 913 1027 1141 1255 1369 1483 1597 1711 1825 1939

1 117 233 349 465 581 697 813 929 1045 1161 1277 1393 1509 1625 1741 1857

1 117 233 349 465 581 697 813 929 1045 1161 1277 1393 1509 1625 1741 1857 0

1 117 233 349 465 581 697 813 929 1045 1161 1277 1393 1509 1625 1741 1857

1 118 235 352 469 586 703 820 937 1054 1171 1288 1405 1522 1639 1756 1873

1 115 229 343 457 571 685 799 913 1027 1141 1255 1369 1483 1597 1711 1825 1939

Figure 4.8 Coefficient of parameters and metabolites concentration: glycerol fermentation plus furmarate (A) Furmarate (B) Ethanol (C) Succinate (D) Glycerol (E) NH3 (F) Formate (G) Biomass (H) Lactate (I) CO2.

(A)

1 90 179 268 357 446 535 624 713 802 891 980 1069 1158 1247 1336 1425 1514 1603 1692 1781 1870

Figure 4.9 Parameters distribution: glycerol anaerobic utilization plus fumarate

EFM1

EFM1 EFM2 EFM3 EFM4 EFM5 EFM6 EFM7 EFM8 EFM9 EFM10 EFM11 EFM12 EFM13 EFM14 EFM15 EFM16 EFM17

. . . . . . . . . . EFM1952

Figure 4.10 EFMs Correlation coefficient: glycerol anaerobic utilization plus fumarate

Figure 4.11 The mode No.320 in glycerol fermentation in medium plus fumarate

The EFMs correlation coefficient about glycerol anaerobic utilization plus fumarate shown in Figure 4.10, that correlation coefficient seems very unanimous blot. The possible reason about EFMs of glycerol fermentation in medium plus fumarate similarity is that fumarate make the reactions of metabolic pathway similar in fumarate related part such as malate convert to fumarate , fumarate convert to succinate , and all of EFMs have succinate as end product.

Finally, analysis of E.coli ferment glycerol that expressed foreign gene dhaB from Klebsiella pneumoniae. In Figure 4.12, the value of coefficient in ethanol and 1,3-propanediol

are higher than others, that explain the major product of recombinant E.coli are ethanol and 1,3-propanediol.becuase of the reactions produce ethanol and 1,3-propanediol that coproduce redox cofactor such as NADH or NAD+. The ratio of NADH and NAD+ are decisive role for cell organization, so intracellular balance about NADH and NAD+ were controlled very careful and attentive.

1 106 211 316 421 526 631 736 841 946 1051 1156 1261 1366 1471 1576 1681

1 104 207 310 413 516 619 722 825 928 1031 1134 1237 1340 1443 1546 1649 1752

1 104 207 310 413 516 619 722 825 928 1031 1134 1237 1340 1443 1546 1649 1752 0

1 104 207 310 413 516 619 722 825 928 1031 1134 1237 1340 1443 1546 1649 1752

1 106 211 316 421 526 631 736 841 946 1051 1156 1261 1366 1471 1576 1681

1 104 207 310 413 516 619 722 825 928 1031 1134 1237 1340 1443 1546 1649 1752

(A)

Figure 4.12 Coefficient of parameters and metabolites concentration: glycerol fermentation plus furmarate (A) 1,3 - PDO (B) Ethanol (C) Succinate (D) Glycerol (E) NH3 (F) Formate (G) Biomass (H)Lactate(I)CO2

1E-13 1E-10 1E-07 0.0001

0.1

1 82 163 244 325 406 487 568 649 730 811 892 973 1054 1135 1216 1297 1378 1459 1540 1621 1702

1 82 163 244 325 406 487 568 649 730 811 892 973 1054 1135 1216 1297 1378 1459 1540 1621 1702 (A)

(B)

Figure 4.13 Parameters distribution: glycerol fermentation express foreign gene

EFM1 EFM2 EFM3 EFM4 EFM5 EFM6 EFM7 EFM8 EFM9 EFM10 EFM11 EFM12 EFM13 EFM14 EFM15 EFM16 EFM17

. . . . . . . . . . . . . . . . . . . EFM1762

Figure 4.14 EFMs Correlation coefficient: glycerol fermentation express foreign gene.

Figure 4.15 The mode No236 in glycerol fermentation expressed foreign gene dhaB.

在文檔中大腸桿菌在無氧環境下利用甘油生產酒精的模擬與分析 (頁 49-62)