Microbial hyaluronic acid production explained

Microbial hyaluronic acid production refers to the process by which microorganisms, such as bacteria and yeast, are utilized in fermentation to synthesize hyaluronic acid (HA).[1] HA is used in a wide range of medical, cosmetic, and biological products because of its high moisture retention and viscoelasticity qualities.[2] HA had originally been extracted from rooster combs in limited quantities.[3] However, challenges such as low yields, high production costs, and ethical issues associated with animal-derived HA has driven the development of microbial production methods for HA.[4]

Although there are other methods for instance chemical synthesis and modification, chemoenzymatic synthesis, enzymatic synthesis; microbial fermentation has been preferred to produce HA because of economical advantages.[5]

Bacterial production

Some bacteria, like Streptococcus, develop an extracellular capsule that contains HA. This capsule functions as a molecular mimic to elude the host's immune system during the infection process in addition to providing adherence and protection.[6] Streptococcus zooepidemicus was used for first commercially HA fermentation, and that is most used bacteria since provides high yields although it is a pathogen microorganism.[7]

Encoding of HA production is carried out by hasA, hasB, hasC, hasD and hasE genes in S. zooepidemicus.[8]

Genes and their functions HA production in S. zooepidemicus !Gene!Enzyme!Function!Reference
hasAHyaluronic acid synthaseHA synthesis and

transport

[9]
hasBUDP-glucose dehydrogenaseUDP-GlcA

biosynthesis

[10] [11]
hasCUDP-glucose pyrophosphorylaseUDP-GlcA

biosynthesis

[12]
hasDAcetyltransferase and

pyrophosphorylase (bifunctional)

UDP-GlcNAc

biosynthesis

[13]
hasEPhosphoglucoisomeraseUDP-GlcNAc

biosynthesis

Genetically modified producers were developed such as Kluysveromyces  lactis,[14]  Lactococcus  lactis,[15] Bacillus  subtilis,[16] Escherichia  coli,[17]   and Corynebacterium glutamicum[18] [19] because of S. zooepidemicus’s pathogeny.

Biological process

Intracellular factors

Metabolism

Intermediates are used from  pathways  essential  to  support cell  growth,  such  as  the  production  of  organic  acids,  polysaccharides during the HA production.[20] HA is not an essential metabolite, and it competes other metabolites to attend the carbon flux in the cell. Reduction potential of S. zooepidemicus may have a role in hyaluronic acid production, because 2 NAD+ are consumed during the synthesis of one monomer. Although NAD+ does not control HA synthesis when NADH oxidase over-expressed,[21] it has a big role in biomass formation.

Some studies showed that balanced intracellular concentration of precursors and their fluxes balanced provides higher molecular weight such as UDP-acetylglucosamine concentration.[22] [23] Enzymes such as hyaluronidase,[24] β-glucuronidase[25] of S. zooepidemicus decrease yield of HA. HA concentration is increased by deletion of associated genes of these enzymes.

On the other hand, some enzymes induce HA production like sucrose-6-phosphatate hydrolase,[26] and hyaluronan synthase.[27] Using combined approaches with these two type enzymes is a good strategy for high yield HA production.

Membrane

HA is produced around the cell, serving as a barrier against the host immune system by the bacteria. Only 8% of HA remains as attached the cell when cells arrived stationary phase. Biosurfactants such as sodium dodecyl sulfate (SDS) are used to gain this product.[28] Hyaluronan synthase, that is a membrane-binding enzyme, is one of the factors that reduces the production of HA. Hyaluronan synthase limits hyaluronic acid production by affecting cell morphology.

Environmental factors

pH

Organic acids formed during HA production by S. zooepidemicus cause pH to decrease Although HA production without pH control is cheaper, it prefers since provides high hyaluronic acid yields..[29] [30]

Temperature

HA production is affected regarding to yield and molecular weight by temperature.[31] HA production increases while bacterial cells are growing above 37°C. However, HA yield decreases while molecular weight is higher with fermentation under 32°C.

Aeration

Although S. zooepidemicus is an aerotolerant anaerobe, hyaluronic acid production is affected by oxygen because NADH/NAD+ balance of cells changes with oxygen amount. Controlling oxygen during the cultivation via agitation rate provides increase both HA yield and molecular weight.[32]

Culture Media Components

The carbon source is one of the media components that has effects on production of microbial HA. Although the glucose[33] [34] is most used one as a carbon source for the HA production; molasses,[35] sucrose,[36] and maltose are used for microbial production.

HA production needs also many amino acids in the culture media therefore nitrogen source concentration has a key.[37]

See also

Notes and References

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  2. Sze JH, Brownlie JC, Love CA . Biotechnological production of hyaluronic acid: a mini review . 3 Biotech . 6 . 1 . 67 . June 2016 . 28330137 . 4754297 . 10.1007/s13205-016-0379-9 .
  3. Ciriminna R, Scurria A, Pagliaro M . 2021 . Microbial production of hyaluronic acid: the case of an emergent technology in the bioeconomy . Biofuels, Bioproducts and Biorefining . en . 15 . 6 . 1604–1610 . 10.1002/bbb.2285 . 1932-104X. free .
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  5. Shikina EV, Kovalevsky RA, Shirkovskaya AI, Toukach PV . Prospective bacterial and fungal sources of hyaluronic acid: A review . Computational and Structural Biotechnology Journal . 20 . 6214–6236 . 2022 . 36420162 . 9676211 . 10.1016/j.csbj.2022.11.013 .
  6. Wessels MR, Moses AE, Goldberg JB, DiCesare TJ . Hyaluronic acid capsule is a virulence factor for mucoid group A streptococci . Proceedings of the National Academy of Sciences of the United States of America . 88 . 19 . 8317–8321 . October 1991 . 1656437 . 52499 . 10.1073/pnas.88.19.8317 . free . 1991PNAS...88.8317W .
  7. Torres-Acosta MA, Castaneda-Aponte HM, Mora-Galvez LM, Gil-Garzon MR, Banda-Magaña MP, Marcellin E, Mayolo-Deloisa K, Licona-Cassani C . Comparative Economic Analysis Between Endogenous and Recombinant Production of Hyaluronic Acid . Frontiers in Bioengineering and Biotechnology . 9 . 680278 . 2021-07-21 . 34368093 . 8334870 . 10.3389/fbioe.2021.680278 . free .
  8. Zhang Y, Luo K, Zhao Q, Qi Z, Nielsen LK, Liu H . Genetic and biochemical characterization of genes involved in hyaluronic acid synthesis in Streptococcus zooepidemicus . Applied Microbiology and Biotechnology . 100 . 8 . 3611–3620 . April 2016 . 26758299 . 10.1007/s00253-016-7286-1 .
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