Appendix A: Abbreviations Used in Metabolic Reactions

Appendix: metabolic network model construction

The main metabolic pathways by which carbon energy sources were used for the production of energy and carbon skeleton needed for the synthesis of intermediary metabolites, building blocks, and end products were considered for streptomyces in a reaction network. In addition to the main catabolic and anabolic pathways, the secondary metabolism involving daptomycin production was included in this network. Furthermore, ammonia and sulfate assimilation reactions were linked to the metabolism. Moreover, in case no specific information for Streptomyces roseosporus was found, corresponding data of closely related streptomyces (e.g., Streptomyces lividans) were included, and data obtained for bacteria (e.g., Escherichia coli) were added when necessary. This reaction network includes Embden-Meyerhof-Parnas pathway (EMP) and pentose phosphate pathways (PPP), tricarboxylic acid cycle (TCA), anaplerotic reactions, ammonia and sulfate assimilation, electron transport reactions, folic acid and thioredoxin reactions. The biosynthesis of aromatic, aspartate, glutamate, pyruvate, and serine family amino acids and histidine, pyrimidine, and purine nucleotides, and the biosynthesis of macromolecular components of biomass such as RNA, DNA, protein, fatty acids, phospholipids, carbohydrate, as well as daptomycin production were added in the network. The anabolic reactions for S. roseosporus for protein, lipid, RNA, DNA, and biomass synthesis were taken from the reported information for Streptomyces coelicolor A3(2), Streptomyces avermitilis and E. coli at the specific growth rate of 0.1 h–1 [1–5].

Although the macromolecular biomass composition changed with the growth rate [5], it was considered constant as the impact of these changes on the intracellular flux distributions was negligible. In this research, a single reaction based on a fixed biomass composition was employed for the formation of biomass throughout the fermentation. The Entener–Doudoroff (ED) pathway in streptomyces was not taken into consideration in the model. The glyoxylate pathway in S. roseosporus was also inactive. The operational P/O ratios for the oxidation processes in S. roseosporus in our simulations were assumed 2 and 1 for NADH and FADH2, respectively. CO2, NH3, in the form of NH4+, and phosphate transporting from the cell to the broth and from the broth to the cell were also assumed to be by passive transport. Furthermore, pathway synthesizing 3-methy-glutamate was also added [6–7].

Appendix A: Abbreviations used in metabolic reactions

Appendix B: Metabolic reactions

Glycolysis Pathway

R1. GLC + ATP → G6P + ADP

R2. G6P ↔ F6P

R3. F6P + ATP ↔ G3P + DHAP + ADP

R4. DHAP ↔ G3P

R5. G3P + PI + NAD + ADP ↔ NADH + 3PG + ATP

R6. 3PG ↔ PEP

R7. PEP + ADP → PYR + ATP

Pentose Phosphate Pathway

R8. G6P + 2 NADP ↔ CO2 + RL5P + 2 NADPH

R9. RL5P ↔ R5P

R10. RL5P ↔ X5P

R11. R5P + X5P ↔ G3P + S7P

R12. X5P + E4P ↔ F6P + G3P

R13. G3P + S7P ↔ E4P + F6P

Branches from Glycolysis Pathway

R14. PYR + NAD + COA → ACCOA + CO2 + NADH

R15. PYR → ACAL + CO2

R16. NAD + ACAL ↔ AC + NADH

R17. PYR + NADH ↔ LAC + NAD

Anaplerotic Reactions

R18. PEP + CO2 + ADP → OA + ATP

R19. PYR + ATP + CO2 → ADP + OA + PI

R20. MAL + NAD → CO2 + NADH + PYR

TCA Cycle

R21. ACCOA + OA → COA + CIT

R22. CIT ↔ ICIT

R23. ICIT + NADP → NADPH + AKG + CO2

R24. AKG + NAD + COA ↔ CO2 + NADH + SUCCOA

R25. GDP + PI + SUCCOA ↔ GTP + SUCC + COA

R26. SUCC + FAD ↔ FUM + FADH2

R27. FUM ↔ MAL

R28. MAL + NAD ↔ NADH + OA

Biosynthesis of Serine Family Amino Acids

R29. 3PG + NAD + GLU → NADH + AKG + PI + SER

R30. THF + SER ↔ GLY + METTHF

R31. SER + ACCOA + H2S → COA + CYS + AC

Biosynthesis of Alanine Family Amino Acids

R32. PYR + NH3 + NADH ↔ ALA + NAD

R33. 2 PYR + NADPH → NADP + OIVAL + CO2

R34. OIVAL + GLU → AKG + VAL

R35. ACCOA + OIVAL + NAD + GLU → COA + NADH + CO2 + AKG + LEU

Biosynthesis of Histidine

R36. R5P + ATP ↔ PRPP + AMP

R37. PRPP + ATP + GLN + 2 NAD → 2 PPI + AKG + AICAR + PI + 2 NADH + HIS

Biosynthesis of Aspartic Family Amino Acids

R38. OA + GLU ↔ ASP + AKG

R39. ASP + ATP + GLN → GLU + ASN + AMP + PPI

R40. ASP + ATP + NADPH → ADP + NADP + PI + ASPSA

R41. ASPSA + NADPH → NADP + HSER

R42. HSER + ATP → ADP + THR + PI

R43. HSER + SUCCOA + CYS ↔ SUCC + HCYS + PYR + NH3 + COA

R44. HCYS + MTHF ↔ THF + MET

R45. THR + PYR + NADPH + GLU → CO2 + NH3 + NADP + AKG + ILE

Biosynthesis of Aromatic Family Amino Acids

R46. E4P + 2 PEP + NADPH + ATP → 4 PI + ADP + CHOR

R47. CHOR + GLN → GLU + PYR + AN

R48. AN + PRPP + SER → PPI + CO2 + G3P + TRP

R49. CHOR → PHEN

R50. PHEN + GLU → CO2 + AKG + PHE

R51. PHEN + NAD + GLU → AKG + TYR + CO2 + NADH

Biosynthesis of Glutamic Family Amino Acids

R52. AKG + NH3 + NADPH ↔ GLU + NADP

R53. GLU + NH3 + ATP → GLN + ADP + PI

R54. GLU + ATP + 2 NADPH + NADH → ADP + 2 NADP + NAD + PI + PRO

R55. ATP + NADPH + 2 GLU → ADP + NADP + PI + AKG + ORN

R56. GLN + 2 ATP + CO2 → GLU + CAP + 2 ADP + PI

R57. ASP + 2 ATP + CAP + NADPH + 2 GLU → ARG + FUM + AKG + AMP + PPI + 2 PI + ADP + NADP

R58. ASPSA + PYR + NADH + NADPH + SUCCOA + GLU → NAD + NADP + COA + AKG + SUCC + MDAPIM

R59. MDAPIM → LYS + CO2

Biosynthesis of D Family Amino Acids

R60. GLU ↔ DGLU

R61. ALA ↔ DALA

R62. SER ↔ DSER

R63. ASN ↔ DASN

Biosynthesis of Nonprotein Amino Acids

R64. ATP + MET → SAM + PPI + PI

R65. SAM + AKG → MAKG + SAH

R66. SAH → HCYS + ADN

R67. MAKG + VAL → MGLU + OIVAL

R68. TRP + O2 → FKYN

R69. FKYN → FOR + KYN

Biosynthesis of Nucleotides

R70. PRPP + 2 GLN + 2 ATP + GLY + FTHF → PPI + 2 GLU + 2 ADP + 2 PI + THF + FGAM

R71. FGAM + 3 ATP + CO2 + ASP → 3 ADP + 3 PI + FUM + AICAR

R72. AICAR + FTHF ↔ THF + IMP

R73. IMP + NAD + 2 ATP + GLN → GLU + AMP + PPI + GDP + NADH + ADP

R74. GDP + ATP ↔ GTP + ADP

R75. GDP + RTHIO + ATP → DGTP + OTHIO + ADP

R76. ADN + ATP ↔ AMP + ADP

R77. AMP + ATP → 2 ADP

R78. ATP + RTHIO → DATP + OTHIO

R79. CAP + ASP + NAD + PRPP → NADH + PPI + UMP + CO2 + PI

R80. ATP + UMP ↔ ADP + UMP

R81. ATP + UDP ↔ ADP + UTP

R82. ATP + UTP + NH3 → ADP + PI + CTP

R83. CDP + ATP ↔ CTP + ADP

R84. CDP + RTHIO + ATP → DCTP + OTHIO + ADP

R85. DCTP + METTHF → DHF + NH3 + DTTP

Folate biosynthesis and Interconversion of One-Carbon Units

R86. DHF + NADPH → NADP + THF

R87. THF + FOR + ATP → FTHF + PI + ADP

R88. FTHF ↔ METHF

R89. METHF + NADPH ↔ METTHF + NADP

R90. METTHF + FADH2 → MTHF + FAD

R91. OTHIO + NADPH → RTHIO + NADP

Biosynthesis of Carbohydrate

R92. F6P + GLN + ACCOA + UTP → GLU + COA + UDPNAG + PPI

R93. G6P + UTP → PPI + UDPGAL

Biosynthesis of Peptidoglycan

R94. UDPNAG + PEP + NADPH → UDPNAM + PI + NADP

R95. 2 DALA + ATP → ALAALA + ADP + PI

Biosynthesis of Triacylglycerol

R96. DHAP + NADH ↔ GL3P + NAD

R97. ACCOA + ATP + CO2 ↔ MALCOA + ADP + PI

R98. MALCOA + ACP → MALACP + COA

R99. ACCOA + ACP → ACACP + COA

R100. ACACP + 6 MALACP + 12 NADPH → 12 NADP + C140ACP + 6 CO2 + 6 ACP

R101. ACACP + 6.5 MALACP + 13 NADPH → 13 NADP + C150ACP + 6.5 CO2 + 6.5 ACP

R102. ACACP + 7 MALACP + 14 NADPH → 14 NADP + C160ACP + 7 CO2 + 7 ACP

R103. ACACP + 7.5 MALACP + 15 NADPH → 15 NADP + C170ACP + 7.5 CO2 + 7.5 ACP

R104. ACACP + 8 MALACP + 15 NADPH → 15 NADP + C181ACP + 8 CO2 + 8 ACP

Biosynthesis of Phospholipid

R105. GL3P + 0.094 C140ACP + 0.294 C150ACP + 0.262 C160ACP + 0.293 C170ACP + 0.057 C181ACP → AGL3P + ACP

R106. AGL3P + 0.094 C140ACP + 0.294 C150ACP + 0.262 C160ACP + 0.293 C170ACP + 0.057 C181ACP → PA + ACP

R107. PA + CTP ↔ CDPDG + PPI

R108. CDPDG + SER ↔ CMP + PS

R109. PS → PE + CO2

R110. CDPDG + GL3P ↔ CMP + PGP

R111. PGP → PI + PG

R112. CDPDG + PG → CMP + CL

Biosynthesis of Teichoic acid

R113. GL3P + CTP → PPI + CDPGL

R114. 12 CDPGL → 12 CMP + POLYGP

Maintenance Energy

R115. ATP → ADP + PI

R116. PPI → 2 PI

R117. NADH + 2 ADP + 2 PI + 0.5 O2 → 2 ATP + NAD

R118. FADH2 + ADP + PI + 0.5 O2 → ATP + FAD

R119. NADPH + NAD → NADP + NADH

R120. SO4 + 2 ATP + 4 NADPH → 2 ADP + H2S + 2 PI + 4 NADP

Biosynthesis of DNA

R121. 0.469 DATP + 1.149 DCTP + 0.469 DTTP + 1.149 DGTP + 4.4 ATP → 4.4 ADP + 4.4 PI + 3.236 PPI + DNA

Biosynthesis of Teichoic acid

R122. 0.518 POLYGP + 0.129 LYS + 0.129 UDPNAG + 0.129 ATP → TEICH + 0.129 UDP + 0.129 ADP + 0.129 PI

Biosynthesis of RNA

R123. 0.600 ATP + 0.826 GTP + 1.031 CTP + 0.662 UTP + 1.25 ATP → 1.25 ADP + 1.25 PI + RNA + 3.119 PPI

Biosynthesis of Peptidoglycan

R124. 1.007 UDPNAM + 1.197 UDPNAG + 1.900 ALAALA + 0.950 ALA + 1.140 MDAPIM + 1.014 DGLU + 0.973 GLY + 5.026 ATP → PEPTIDOGLYCAN + 0.950 DALA + 1.197 UDP + 1.007 UMP + 5.026 ADP + 5.026 PI

Biosynthesis of Phospholipid

R125. 1.089 PE + 0.250 PG + 0.052 CL → PHOSPHOLIPID

Biosynthesis of Triacylglycerol

R126. 1.244 GL3P + 0.050 C140ACP + 1.677 C150ACP + 0.421 C160ACP + 1.570 C170ACP + 0.014 C181ACP → TAG + 3.732 ACP + 1.244 PI

Biosynthesis of Protein

R127. 1.350 ALA + 0.352 ARG + 0.394 ASN + 0.391 ASP + 0.165 CYS + 0.382 GLN + 0.379 GLU + 2.015 GLY + 0.131 HIS + 0.477 ILE + 0.742 LEU + 0.499 LYS + 0.221 MET + 0.238 PHE + 0.422 PRO + 0.459 SER + 0.465 THR + 0.059 TRP + 0.159 TYR + 0.797 VAL + 40.0 ATP → 40.0 ADP + 40.0 PI + PROTEIN

Biosynthesis of Carbohydrate

R128. 1.897 UDPNAG + 3.794 UDPGAL → 5.691 UDP + CARBOHYDRATE

Biomass Synthesis

R129. 0.412 PROTEIN + 0.167 RNA + 0.036 DNA + 0.042 PHOSPHOLIPID + 0.033 TAG + 0.110 PEPTIDOGLYCAN + 0.044 CARBOHYDRATE + 0.066 TEICH + 47 ATP → BIOMASS + 47 ADP + 47 PI

Daptomycin Synthesis

R130. DEC + DALA + 2 GLY + 3 ASP + DASN + ORN + MGLU + DSER + THR + TRP + KYN + 14 ATP → DAPTOMYCIN + 14 ADP

References

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