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Applied Microbiology and Biotechnology

, Volume 103, Issue 17, pp 6989–7001 | Cite as

Enhancement of acetyl-CoA by acetate co-utilization in recombinant Lactococcus lactis cultures enables the production of high molecular weight hyaluronic acid

  • Kirubhakaran Puvendran
  • Guhan JayaramanEmail author
Biotechnological products and process engineering
  • 147 Downloads

Abstract

The molecular weight of hyaluronic acid (HA) is a critical property which determines its usage in various biomedical applications. This study investigates the correlation between the availability of a critical cofactor, acetyl-CoA, the concentration of a limiting precursor, UDP-N-acetylglucosamine (UDP-GlcNAc), and the molecular weight of HA (MWHA) produced by recombinant Lactococcus lactis MKG6 cultures. This strain expressed three heterologous HA-pathway genes obtained from the has operon of Streptococcus zooepidemicus in an ldh-mutant host strain, L. lactis NZ9020. A flux balance analysis, performed using the L. lactis genome-scale metabolic network, showed a positive correlation of acetyl-CoA flux with the UDP-GlcNAc flux and the experimental data on HA productivity. To increase the intracellular levels of acetyl-CoA, acetate was supplemented as a pulse feed in anaerobic batch cultures. However, acetate is effectively utilized only in the presence of glucose and exhaustion of glucose resulted in decreasing the final MWHA (1.5 MDa). Co-supplementation of acetate resulted in enhancing the acetyl-CoA and UDP-GlcNAc levels as well as the MWHA to 2.5 MDa. This logic was extended to fed-batch cultures, designed with a pH-based feedback control of glucose feeding and pulse acetate supplementation. When the glucose feed concentration was optimally adjusted to prevent glucose exhaustion or accumulation, the acetate utilization was found to be high, resulting in significantly enhanced levels of acetyl-CoA and UDP-GlcNAc as well as a MWHA of 3.4 MDa, which was sustained at this value throughout the process. This study provides the possibility of commercially producing high MWHA using recombinant L. lactis strains.

Keywords

Acetyl-CoA Fed-batch process Hyaluronic acid Molecular weight Lactococcus lactis 

Notes

Acknowledgments

The authors would like to acknowledge the support provided by the Ministry of Human Resources Development (Govt. of India), for fellowship to Kirubhakaran Puvendran. We would also like to acknowledge the Department of Chemistry, IIT Madras, for helping us with NMR spectroscopy and thermo-gravimetric analysis.

Funding

This work was financially supported by the Department of Biotechnology (Ministry of Science and Technology, Govt. of India) through Project No. BT/PR13815/BBE/117/61/2015.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.

Supplementary material

253_2019_9987_MOESM1_ESM.pdf (766 kb)
ESM 1 (PDF 766 kb)

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Biotechnology, Bhupat and Jyoti Mehta School of BiosciencesIndian Institute of Technology MadrasChennaiIndia
  2. 2.Bioprocess and Metabolic Engineering Laboratory, Department of Biotechnology, Bhupat and Jyoti Mehta School of BiosciencesIndian Institute of Technology MadrasChennaiIndia

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