Human FAS ELISA Matched Antibody Pair

Camelina sativais an emerging biotechnology oil crop. However, more information is needed regarding its innate lipid enzyme specificities. We have therefore characterized several triacylglycerol (TAG) producing enzymes by measuringin vitrosubstrate specificities using different combinations of acyl-acceptors (diacylglycerol, DAG) and donors. Specifically,C. sativaacyl-CoA:diacylglycerol acyltransferase (DGAT) 1 and 2 (which both use acyl-CoA as acyl donor) and phospholipid:diacylglycerol acyltransferase (PDAT, with phosphatidylcoline as acyl donor) were studied. The results show that the DGAT1 and DGAT2 specificities are complementary, with DGAT2 exhibiting a high specificity for acyl acceptors containing only polyunsaturated fatty acids (FAs), whereas DGAT1 prefers acyl donors with saturated and monounsaturated FAs. Furthermore, the combination of substrates that resulted in the highest activity for DGAT2, but very low activity for DGAT1, corresponds to TAG species previously shown to increase inC. sativaseeds with downregulated DGAT1. Similarly, the combinations of substrates that gave the highest PDAT1 activity were also those that produce the two TAG species (54:7 and 54:8 TAG) with the highest increase in PDAT overexpressingC. sativaseeds. Thus, thein vitrodata correlate well with the changes in the overall fatty acid profile and TAG species inC. sativaseeds with altered DGAT1 and PDAT activity. Additionally,in vitrostudies ofC. sativaphosphatidycholine:diacylglycerol cholinephosphotransferase (PDCT), another activity involved in TAG biosynthesis, revealed that PDCT accepts substrates with different desaturation levels. Furthermore, PDCT was unable to use DAG with ricineoleyl groups, and the presence of this substrate also inhibited PDCT from using other DAG-moieties. This gives insights relating to previousin vivostudies regarding this enzyme.

Background Schizochytriumspecies are known for their abundant production of docosahexaenoic acid (DHA). Low temperatures can promote the biosynthesis of polyunsaturated fatty acids (PUFAs) in many species. This study investigates low-temperature effects on DHA biosynthesis inSchizochytriumsp. TIO01 and its underlying mechanism. Results TheSchizochytriumfatty acid biosynthesis pathway was evaluated based on de novo genome assembly (contig N50 = 2.86 Mb) and iTRAQ-based protein identification. Our findings revealed that desaturases, involved in DHA synthesis via the fatty acid synthase (FAS) pathway, were completely absent. The polyketide synthase (PKS) pathway and the FAS pathway are, respectively, responsible for DHA and saturated fatty acid synthesis inSchizochytrium. Analysis of fatty acid composition profiles indicates that low temperature has a significant impact on the production of DHA inSchizochytrium, increasing the DHA content from 43 to 65% of total fatty acids. However, the expression levels of PKS pathway genes were not significantly regulated as the DHA content increased. Further, gene expression analysis showed that pathways related to the production of substrates (acetyl-CoA and NADPH) for fatty acid synthesis (the branched-chain amino acid degradation pathway and the pentose phosphate pathway) and genes related to saturated fatty acid biosynthesis (the FAS pathway genes and malic enzyme) were, respectively, upregulated and downregulated. These results indicate that low temperatures increase the DHA content by likely promoting the entry of relatively large amounts of substrates into the PKS pathway. Conclusions In this study, we provide genomic, proteomic, and transcriptomic evidence for the fatty acid synthesis pathway inSchizochytriumand propose a mechanism by which low temperatures promote the accumulation of DHA inSchizochytrium. The high-quality and nearly complete genome sequence ofSchizochytriumprovides a valuable reference for investigating the regulation of polyunsaturated fatty acid biosynthesis and the evolutionary characteristics ofThraustochytriidaespecies.