Some plant trans-1,4-prenyltransferases (TPTs) produce ultrahigh molecular weight trans-1,4-polyisoprene (TPI) with a molecular weight of over 1.0 million. Although plant-derived TPI has been utilized in various industries, its biosynthesis and physiological function(s) are unclear. Here, we identified three novel Eucommia ulmoides TPT isoforms—EuTPT1, 3, and 5, which synthesized TPI in vitro without other components. Crystal structure analysis of EuTPT3 revealed a dimeric architecture with a central hydrophobic tunnel. Mutation of Cys94 and Ala95 on the central hydrophobic tunnel no longer synthesizd TPI, indicating that Cys94 and Ala95 were essential for forming the dimeric architecture of ultralong-chain TPTs and TPI biosynthesis. A spatiotemporal analysis of the physiological function of TPI in E. ulmoides suggested that it is involved in seed development and maturation. Thus, our analysis provides functional and mechanistic insights into TPI biosynthesis and uncovers biological roles of TPI in plants.

一些植物反式-1,4-异戊二烯基转移酶（TPT）产生分子量超过100万的超高分子量反式-1,4-聚异戊二烯（TPI）。尽管植物来源的 TPI 已被用于各个行业，但其生物合成和生理功能尚不清楚。在这里，我们确定了三种新型杜仲TPT 异构体-EuTPT1、3 和 5，它们在没有其他成分的情况下在体外合成了 TPI。EuTPT3 的晶体结构分析揭示了具有中心疏水隧道的二聚体结构。Cys94 和 Ala95 在中央疏水通道上的突变不再合成 TPI，表明 Cys94 和 Ala95 对于形成超长链 TPT 和 TPI 生物合成的二聚体结构至关重要。对E. ulmoides中 TPI 生理功能的时空分析表明它参与种子发育和成熟。因此，我们的分析提供了对 TPI 生物合成的功能和机制见解，并揭示了 TPI 在植物中的生物学作用。