TY - JOUR
T1 - Increased 3′-Phosphoadenosine-5′-phosphosulfate Levels in Engineered Escherichia coli Cell Lysate Facilitate the In Vitro Synthesis of Chondroitin Sulfate A
AU - Badri, Abinaya
AU - Williams, Asher
AU - Xia, Ke
AU - Linhardt, Robert J.
AU - Koffas, Mattheos A.G.
N1 - Publisher Copyright:
© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
PY - 2019/9/1
Y1 - 2019/9/1
N2 - Chondroitin sulfates (CSs) are linear glycosaminoglycans that have important applications in the medical and food industries. Engineering bacteria for the microbial production of CS will facilitate a one-step, scalable production with good control over sulfation levels and positions in contrast to extraction from animal sources. To achieve this goal, Escherichia coli (E. coli) is engineered in this study using traditional metabolic engineering approaches to accumulate 3′-phosphoadenosine-5′-phosphosulfate (PAPS), the universal sulfate donor. PAPS is one of the least-explored components required for the biosynthesis of CS. The resulting engineered E. coli strain shows an ≈1000-fold increase in intracellular PAPS concentrations. This study also reports, for the first time, in vitro biotransformation of CS using PAPS, chondroitin, and chondroitin-4-sulfotransferase (C4ST), all synthesized from different engineered E. coli strains. A 10.4-fold increase is observed in the amount of CS produced by biotransformation by employing PAPS from the engineered PAPS-accumulating strain. The data from the biotransformation experiments also help evaluate the reaction components that need improved production to achieve a one-step microbial synthesis of CS. This will provide a new platform to produce CS.
AB - Chondroitin sulfates (CSs) are linear glycosaminoglycans that have important applications in the medical and food industries. Engineering bacteria for the microbial production of CS will facilitate a one-step, scalable production with good control over sulfation levels and positions in contrast to extraction from animal sources. To achieve this goal, Escherichia coli (E. coli) is engineered in this study using traditional metabolic engineering approaches to accumulate 3′-phosphoadenosine-5′-phosphosulfate (PAPS), the universal sulfate donor. PAPS is one of the least-explored components required for the biosynthesis of CS. The resulting engineered E. coli strain shows an ≈1000-fold increase in intracellular PAPS concentrations. This study also reports, for the first time, in vitro biotransformation of CS using PAPS, chondroitin, and chondroitin-4-sulfotransferase (C4ST), all synthesized from different engineered E. coli strains. A 10.4-fold increase is observed in the amount of CS produced by biotransformation by employing PAPS from the engineered PAPS-accumulating strain. The data from the biotransformation experiments also help evaluate the reaction components that need improved production to achieve a one-step microbial synthesis of CS. This will provide a new platform to produce CS.
KW - 3′-phosphoadenosine-5′-phosphosulfate reductases
KW - 3′-phosphoadenosine-5′-phosphosulfates
KW - Escherichia coli
KW - chondroitin sulfates
KW - metabolic engineering
UR - http://www.scopus.com/inward/record.url?scp=85071782512&partnerID=8YFLogxK
U2 - 10.1002/biot.201800436
DO - 10.1002/biot.201800436
M3 - Article
C2 - 31180182
AN - SCOPUS:85071782512
SN - 1860-6768
VL - 14
JO - Biotechnology Journal
JF - Biotechnology Journal
IS - 9
M1 - 1800436
ER -