Zienkiewicz Krzysztof, Zienkiewicz Agnieszka, Poliner Eric, Du Zhi-Yan, Vollheyde Katharina, Herrfurth Cornelia, Marmon Sofia, Farré Eva M, Feussner Ivo, Benning Christoph
Michigan State University-US Department of Energy Plant Research Laboratory, Michigan State University, East Lansing, MI 48824 USA ; Department of Plant Biochemistry, Albrecht-von-Haller-Institute for Plant Sciences, Georg-August-University, 37073 Göttingen, Germany.
Michigan State University-US Department of Energy Plant Research Laboratory, Michigan State University, East Lansing, MI 48824 USA ; Department of Plant Biochemistry, Albrecht-von-Haller-Institute for Plant Sciences, Georg-August-University, 37073 Göttingen, Germany ; Great Lakes Bioenergy Center, Michigan State University, East Lansing, MI 48824 USA.
Biotechnol Biofuels. 2017 Jan 3;10:8. doi: 10.1186/s13068-016-0686-8. eCollection 2017.
Photosynthetic microalgae are considered a viable and sustainable resource for biofuel feedstocks, because they can produce higher biomass per land area than plants and can be grown on non-arable land. Among many microalgae considered for biofuel production, (CCMP1779) is particularly promising, because following nutrient deprivation it produces very high amounts of triacylglycerols (TAG). The committed step in TAG synthesis is catalyzed by acyl-CoA:diacylglycerol acyltransferase (DGAT). Remarkably, a total of 13 putative DGAT-encoding genes have been previously identified in CCMP1779 but most have not yet been studied in detail.
Based on their expression profile, six out of 12 type-2 DGAT-encoding genes (-) were chosen for their possible role in TAG biosynthesis and the respective cDNAs were expressed in a TAG synthesis-deficient mutant of yeast. Yeast expressing accumulated TAG to the highest level. Over-expression of in CCMP1779 grown in N-replete medium resulted in levels of TAG normally observed only after N deprivation. Reduced growth rates accompanied over-expression in CCMP1779. Constitutive expression of in was accompanied by increased TAG content in seeds and leaves. A broad substrate specificity for NoDGTT5 was revealed, with preference for unsaturated acyl groups. Furthermore, NoDGTT5 was able to successfully rescue the Arabidopsis - mutant by restoring the TAG content in seeds.
Taken together, our results identified as the most promising gene for the engineering of TAG synthesis in multiple hosts among the 13 DGAT-encoding genes of CCMP1779. Consequently, this study demonstrates the potential of NoDGTT5 as a tool for enhancing the energy density in biomass by increasing TAG content in transgenic crops used for biofuel production.
光合微藻被认为是生物燃料原料的一种可行且可持续的资源，因为它们每单位土地面积能产生比植物更高的生物量，并且可以在非耕地上生长。在众多被考虑用于生物燃料生产的微藻中，（CCMP1779）特别有前景，因为在营养缺乏后它能产生非常大量的三酰甘油（TAG）。TAG合成中的关键步骤由酰基辅酶A：二酰甘油酰基转移酶（DGAT）催化。值得注意的是，此前在CCMP1779中总共鉴定出13个假定的DGAT编码基因，但大多数尚未进行详细研究。
基于它们的表达谱，从12个2型DGAT编码基因（-）中选择了6个，因其在TAG生物合成中可能发挥的作用，并且各自的cDNA在酵母的TAG合成缺陷突变体中表达。表达的酵母积累TAG的水平最高。在氮充足培养基中生长的CCMP1779中过表达，导致TAG水平通常仅在氮缺乏后才观察到。CCMP1779中过表达伴随着生长速率降低。在中组成型表达伴随着种子和叶片中TAG含量增加。揭示了NoDGTT5具有广泛的底物特异性，偏好不饱和酰基。此外，NoDGTT5能够通过恢复种子中的TAG含量成功拯救拟南芥-突变体。
综上所述，我们的结果确定在CCMP1779的13个DGAT编码基因中，是在多个宿主中进行TAG合成工程最有前景的基因。因此，本研究证明了NoDGTT5作为一种工具的潜力，可通过增加用于生物燃料生产的转基因作物中的TAG含量来提高生物质中的能量密度。
