天然紫外吸收剂gadusol在毕赤酵母中的生物合成及其性能分析

Synthesis and properties of the natural ultraviolet absorber gadusol in Komagataella phaffii

摘要:

Gadusol是一种具有抗氧化能力的高效天然紫外吸收物质，广泛存在于微生物、藻类及鱼卵等水生生物中。为解决其天然提取量低且环境不友好等问题，本研究将斑马鱼来源的gadusol合成途径引入毕赤酵母，成功构建了能够合成gadusol的重组毕赤酵母，进一步过表达了来源于树干毕赤酵母的木糖同化基因以提高其关键底物景天庚酮糖-7-磷酸的含量。结果表明木糖的利用是提高gadusol产量的有效策略，在纯木糖培养基中，gadusol的产量达到141.8 mg/L (32.3 mg/g 细胞干重)，约是在纯葡萄糖培养基中的46倍。产物在275-305 nm范围内存在明显吸收，λmax=290 nm。同时，该物质具有一定的抗氧化能力，在反应5 h后，铁离子还原抗氧化能力(ferric ion reducing antioxidant power, FRAP)、2,2′-联氮双(3-乙基苯并噻唑啉-6-磺酸)二铵盐[2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid), ABTS]和1,1-二苯基-2-苦基肼自由基(1,1-diphenyl-2-picrylhydrazyl, DPPH)结果分别出现了90.83%、25.50%和131.80%的变化率，推测其可能作为一种长效的抗氧化剂。本研究证明了毕赤酵母作为底盘生物合成天然紫外吸收剂的巨大潜能，生物合成的gadusol具有良好的紫外吸收与抗氧化性能，为gadusol的工业化生产及应用提供了理论基础。

Abstract:

Gadusol, an efficient natural ultraviolet (UV) absorbing substance with antioxidant capacity, is ubiquitous in aquatic organisms such as microorganisms, algae, and fish eggs. In order to address issues such as its low natural extraction yield and environmental unfriendliness, we introduced the gadusol synthesis pathway from zebrafish into Komagataella phaffii and successfully constructed the recombinant strain capable of synthesizing gadusol. The xylose assimilation genes derived from Scheffersomyces stipitis were further introduced into the recombinant strain to increase the content of the key substrate sedoheptulose 7-phosphate (S7P). The results showed that the utilization of xylose was an effective strategy to increase the yield of gadusol. In the medium with only xylose as the substrate, the yield of gadusol reached 141.8 mg/L (32.3 mg/g dry cell weight, DCW), which was about 46 times of that in the medium with only glucose as the substrate. The product showed obvious absorption in the range of 275–305 nm, with the maximum absorption at 290 nm. Moreover, the product demonstrated antioxidant capacity. After reaction for 5 h, the ferric ion reducing antioxidant power (FRAP), 2,2′-azino- bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging rates changed by 90.83%, 25.50%, and 131.80%, respectively, which suggested that the product may be used as a long-acting antioxidant. This study demonstrated the great potential of K. phaffii as a chassis for the biosynthesis of natural UV absorbers. The biosynthesized gadusol has good UV absorption and antioxidant properties, which provides a theoretical basis for the industrial production and application of gadusol.