生物技术通报 ›› 2022, Vol. 38 ›› Issue (2): 252-262.doi: 10.13560/j.cnki.biotech.bull.1985.2021-0443
收稿日期:
2021-04-06
出版日期:
2022-02-26
发布日期:
2022-03-09
作者简介:
马艳琴,女,硕士研究生,研究方向:透明质酸的生物合成;E-mail: 基金资助:
MA Yan-qin1(), QIU Yi-bin2(), LI Sha1(), XU Hong1
Received:
2021-04-06
Published:
2022-02-26
Online:
2022-03-09
摘要:
透明质酸(hyaluronic acid,HA)是广泛存在于生物体内的功能性糖胺高分子聚合物,在日化、医疗和食品领域应用前景广阔。随着基因工程与代谢工程等合成生物学技术的发展,人们对HA的生物合成过程和机理解析越发深入的同时,也伴随一些新的挑战来临。该综述从分子生物学角度总结了HA的关键合成酶基因及合成途径,对不同来源的HA合成酶进行了氨基酸序列比对分析,并详细描述了HA在常用微生物宿主中生产的安全性、底物不平衡性、发酵低溶氧性等瓶颈问题及其相应的合成生物学策略开发,从而归纳出了目前微生物中高产HA的有效策略,为进一步推动HA的绿色生物制造提供了重要的科学依据和理论支撑。
马艳琴, 邱益彬, 李莎, 徐虹. 透明质酸的生物合成及其代谢工程的研究进展[J]. 生物技术通报, 2022, 38(2): 252-262.
MA Yan-qin, QIU Yi-bin, LI Sha, XU Hong. Research Progress in the Biosynthesis and Metabolic Engineering of Hyaluronic Acid[J]. Biotechnology Bulletin, 2022, 38(2): 252-262.
Microorganism | Strategy | HA /(g·L-1) | Molecular weight/Da | Reference |
---|---|---|---|---|
S. zooepidemicus CKD 117 | Optimizing fermentation conditions | 5.4 | 2.9×106 | [ |
S. zooepidemicus WSH-24 | Intermittent alkaline-stress strategy | 6.5 | - | [5] |
S. zooepidemicus ATCC 39920 | Using oxygen vector and optimum impeller tip speed | 4.25 | 1.54×107 | [ |
S. zooepidemicus BCRC15414 | A two-stage anoxic(static)-aerobic sequential culture | 0.51 | 3.0×103 | [ |
S. zooepidemicus ATCC 39920 | Redirection of carbon flux by addition of glycolytic inhibitors,glutamine and iodoacetate | 5 | 3.1-3.2×106 | [ |
S. zooepidemicus ATCC 39920 | Overexpression of scrB in S. zooepidemicus ΔfruK | 5.6 | - | [ |
S. zooepidemicus IBRC-M 10919 Δhyl | Removal of the hyaluronidase-encoding gene | 6 | 3.8×106 | [ |
S. zooepidemicus HA-13-06 | Two-stage fermentation process | 4.75 | 2.36×106 | [ |
S. zooepidemicus MTCC 3523 | Dissolved oxygen and N-acetyl glucosamine supply | 2.4 | 2.53×106 | [ |
S. thermophilus YIT 2084 | Fermentation conditions and soybean peptide supplementation | 0.208 | 1.3×105 | [ |
S. thermophilus YIT 2084 | Co-expressing HA synthase hasA and hasB | 1.2 | 1×106 | [4] |
S. equisimilis mutant nc2168 | Optimization of flask culture medium and conditions | 0.225 | - | [ |
S. iniae | Ultraviolet mutation | 0.12 | 3×105 | [ |
S. sp. ID9102 | Optimization of medium components | 6.94 | 5.9×106 | [ |
E. coli Top10 | Co-expressing HA synthase hasA and ugd from S. equisimilis | 0.19 | 3.5×105-1.9×106 | [ |
E. coli BL21 | Expression of hyaluronan synthase hasA from S. zooepidemicus ATCC 39920 | 0.532 | 3.46×104 | [ |
B. subtilis 168 | Inducible expression of pmHAS,tuaD and gtaB | 6.8 | 4.55×106 | [ |
Coexpression committed genes(hasA,tuaD,gtaB,glmU,glmM,and glmS);downregulate pfkA expression of the glycolytic pathway;integration of LHAase with the activity at 1.62×106 U/mL | 19.38 | 6.62×103 | [ | |
Engineering of cell membrane by overexpressing pgsA,clsA and reducing the expression of ftsZ | 1.25 | 2.1×106 | [ | |
B. subtilis WB600 | Regulation of temperature using the recombinant strain carrying hasA gene from S. ubris,hasB,and hasC from B. subtilis WB600 | 3.65 | 0.39-6.94×106 | [ |
C. glutamicum 13032 | Coexpression gene hasA from S. equisimilis,hasB from C. glutamicum ATCC13032 | 8.3 | 1.3×106 | [ |
Coexpression gene hasA from S. equisimilis,hasB from C. glutamicum ATCC13032;deletion of lactate dehydrogenase | 21.6 | 1.28×106 | [ | |
An additional promoter PdapB for driving hasB expression;deletion of fba,zwf deletion and knockout of lactate/acetate pathway | 28.7 | 0.21×106 | [ | |
Overexpression of hyaluronan synthase spHasA,enzymes of intermediate metabolic pathways and attenuation of polysaccharide biosynthesis;Disruption of the encapsulation with the leech hyaluronidase | 34.2-74.1 | 5.3×104-3.31×105 | [ | |
Streptomyces albulus | Overexpression of gene hasA from S. zooepidemicus,gene udgA,glmU,and gtaB genes from Streptomyces avermitilis | 6.2 | 2×106 | [ |
Pichia pastoris | Overexpression of xhasA2 and xhasB genes from Xenopus laevis;hasC,hasD,and hasE genes from P. pastoris | 0.8-1.7 | 1.2-2.5×106 | [ |
Lactococcus lactis CES15 | Expression of hasA gene under the control of the PnisA promoter | 6.09 | - | [ |
表1 不同工程菌生产透明质酸的产量
Table 1 Yield and molecular weight of hyaluronic acid produced by different hosts
Microorganism | Strategy | HA /(g·L-1) | Molecular weight/Da | Reference |
---|---|---|---|---|
S. zooepidemicus CKD 117 | Optimizing fermentation conditions | 5.4 | 2.9×106 | [ |
S. zooepidemicus WSH-24 | Intermittent alkaline-stress strategy | 6.5 | - | [5] |
S. zooepidemicus ATCC 39920 | Using oxygen vector and optimum impeller tip speed | 4.25 | 1.54×107 | [ |
S. zooepidemicus BCRC15414 | A two-stage anoxic(static)-aerobic sequential culture | 0.51 | 3.0×103 | [ |
S. zooepidemicus ATCC 39920 | Redirection of carbon flux by addition of glycolytic inhibitors,glutamine and iodoacetate | 5 | 3.1-3.2×106 | [ |
S. zooepidemicus ATCC 39920 | Overexpression of scrB in S. zooepidemicus ΔfruK | 5.6 | - | [ |
S. zooepidemicus IBRC-M 10919 Δhyl | Removal of the hyaluronidase-encoding gene | 6 | 3.8×106 | [ |
S. zooepidemicus HA-13-06 | Two-stage fermentation process | 4.75 | 2.36×106 | [ |
S. zooepidemicus MTCC 3523 | Dissolved oxygen and N-acetyl glucosamine supply | 2.4 | 2.53×106 | [ |
S. thermophilus YIT 2084 | Fermentation conditions and soybean peptide supplementation | 0.208 | 1.3×105 | [ |
S. thermophilus YIT 2084 | Co-expressing HA synthase hasA and hasB | 1.2 | 1×106 | [4] |
S. equisimilis mutant nc2168 | Optimization of flask culture medium and conditions | 0.225 | - | [ |
S. iniae | Ultraviolet mutation | 0.12 | 3×105 | [ |
S. sp. ID9102 | Optimization of medium components | 6.94 | 5.9×106 | [ |
E. coli Top10 | Co-expressing HA synthase hasA and ugd from S. equisimilis | 0.19 | 3.5×105-1.9×106 | [ |
E. coli BL21 | Expression of hyaluronan synthase hasA from S. zooepidemicus ATCC 39920 | 0.532 | 3.46×104 | [ |
B. subtilis 168 | Inducible expression of pmHAS,tuaD and gtaB | 6.8 | 4.55×106 | [ |
Coexpression committed genes(hasA,tuaD,gtaB,glmU,glmM,and glmS);downregulate pfkA expression of the glycolytic pathway;integration of LHAase with the activity at 1.62×106 U/mL | 19.38 | 6.62×103 | [ | |
Engineering of cell membrane by overexpressing pgsA,clsA and reducing the expression of ftsZ | 1.25 | 2.1×106 | [ | |
B. subtilis WB600 | Regulation of temperature using the recombinant strain carrying hasA gene from S. ubris,hasB,and hasC from B. subtilis WB600 | 3.65 | 0.39-6.94×106 | [ |
C. glutamicum 13032 | Coexpression gene hasA from S. equisimilis,hasB from C. glutamicum ATCC13032 | 8.3 | 1.3×106 | [ |
Coexpression gene hasA from S. equisimilis,hasB from C. glutamicum ATCC13032;deletion of lactate dehydrogenase | 21.6 | 1.28×106 | [ | |
An additional promoter PdapB for driving hasB expression;deletion of fba,zwf deletion and knockout of lactate/acetate pathway | 28.7 | 0.21×106 | [ | |
Overexpression of hyaluronan synthase spHasA,enzymes of intermediate metabolic pathways and attenuation of polysaccharide biosynthesis;Disruption of the encapsulation with the leech hyaluronidase | 34.2-74.1 | 5.3×104-3.31×105 | [ | |
Streptomyces albulus | Overexpression of gene hasA from S. zooepidemicus,gene udgA,glmU,and gtaB genes from Streptomyces avermitilis | 6.2 | 2×106 | [ |
Pichia pastoris | Overexpression of xhasA2 and xhasB genes from Xenopus laevis;hasC,hasD,and hasE genes from P. pastoris | 0.8-1.7 | 1.2-2.5×106 | [ |
Lactococcus lactis CES15 | Expression of hasA gene under the control of the PnisA promoter | 6.09 | - | [ |
Field | Application | HA molecular weight/Da | Reference |
---|---|---|---|
Medicine | Treatment of inflam- matory skin diseases. | 6×106 | [70-72] |
Anticancer and anti- proliferative | 1.2×105 | [73-74] | |
Treatment of inflam- matory joint diseases | 1×106 | [75-77] | |
Anti-diabetic | - | [ | |
Immunomodulatory | 2×104 | [ | |
Cosmetics | Anti-aging | <1×104 | [ |
Skin-repairing | 1-10×104 | [ | |
Food | Food and health | <1×104 | [ |
表2 透明质酸应用及对应分子量总结
Table 2 Summary of hyaluronic acid applications and corresponding molecular weights
Field | Application | HA molecular weight/Da | Reference |
---|---|---|---|
Medicine | Treatment of inflam- matory skin diseases. | 6×106 | [70-72] |
Anticancer and anti- proliferative | 1.2×105 | [73-74] | |
Treatment of inflam- matory joint diseases | 1×106 | [75-77] | |
Anti-diabetic | - | [ | |
Immunomodulatory | 2×104 | [ | |
Cosmetics | Anti-aging | <1×104 | [ |
Skin-repairing | 1-10×104 | [ | |
Food | Food and health | <1×104 | [ |
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