title: Analysis of triglyceride synthesis unveils a green algal soluble diacylglycerol acyltransferase and provides clues to potential enzymatic components of the chloroplast pathway.
abstract: Microalgal triglyceride (TAG) synthesis has attracted considerable attention. Particular emphasis has been put towards characterizing the algal homologs of the canonical rate-limiting enzymes, diacylglycerol acyltransferase (DGAT) and phospholipid:diacylglycerol acyltransferase (PDAT). Less work has been done to analyze homologs from a phylogenetic perspective. In this work, we used HMMER iterative profiling and phylogenetic and functional analyses to determine the number and sequence characteristics of algal DGAT and PDAT, as well as related sequences that constitute their corresponding superfamilies. We included most algae with available genomes, as well as representative eukaryotic and prokaryotic species.
  
  
  
  Amongst our main findings, we identified a novel clade of DGAT1-like proteins exclusive to red algae and glaucophyta and a previously uncharacterized subclade of DGAT2 proteins with an unusual number of transmembrane segments. Our analysis also revealed the existence of a novel DGAT exclusive to green algae with moderate similarity to plant soluble DGAT3. The DGAT3 clade shares a most recent ancestor with a group of uncharacterized proteins from cyanobacteria. Subcellular targeting prediction suggests that most green algal DGAT3 proteins are imported to the chloroplast, evidencing that the green algal chloroplast might have a soluble pathway for the de novo synthesis of TAGs. Heterologous expression of C. reinhardtii DGAT3 produces an increase in the accumulation of TAG, as evidenced by thin layer chromatography.
  
  
  
  Our analysis contributes to advance in the knowledge of complex superfamilies involved in lipid metabolism and provides clues to possible enzymatic players of chloroplast TAG synthesis.
authors.affiliation: Instituto de Energía y Desarrollo Sustentable, Comisión Nacional de Energía Atómica, Centro Atómico Bariloche, Av. Bustillo 9500, 8400S. C. de Bariloche, Río Negro, Argentina.
authors.forename: Carolina
authors.initials: C
authors.lastname: Bagnato
authors.affiliation: Instituto de Energía y Desarrollo Sustentable, Comisión Nacional de Energía Atómica, Centro Atómico Bariloche, Av. Bustillo 9500, 8400S. C. de Bariloche, Río Negro, Argentina.
authors.forename: María B
authors.initials: MB
authors.lastname: Prados
authors.affiliation: Instituto de Investigaciones Bioquímicas de La Plata (INIBIOLP-CONICET-UNLP), Facultad de Ciencias Médicas, Universidad Nacional de La Plata, Calle 60 y 120 s/n, 1900, La Plata, Argentina.
authors.forename: Gisela R
authors.initials: GR
authors.lastname: Franchini
authors.affiliation: Instituto de Investigaciones Bioquímicas de La Plata (INIBIOLP-CONICET-UNLP), Facultad de Ciencias Médicas, Universidad Nacional de La Plata, Calle 60 y 120 s/n, 1900, La Plata, Argentina.
authors.forename: Natalia
authors.initials: N
authors.lastname: Scaglia
authors.affiliation: Universidad de Buenos Aires. CONICET Instituto de Investigaciones Cardiológicas (ININCA), - Laboratorio de Glico-Inmuno-Biología, Marcelo T. de Alvear 2270, C1122AAJ, Buenos Aires, Argentina.
authors.forename: Silvia E
authors.initials: SE
authors.lastname: Miranda
authors.affiliation: Instituto de Investigaciones Biológicas (IIB-CONICET-UNMdP), Facultad de Ciencias Exactas y Naturales, Universidad Nacional de Mar del Plata, CC 1245, 7600, Mar del Plata, Argentina. mvbeligni@mdp.edu.ar.
authors.forename: María V
authors.initials: MV
authors.lastname: Beligni
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000460:Chlorophyta / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D002736:Chloroplasts / Q000235:genetics / Q000378:metabolism*; D019295:Computational Biology / Q000379:methods; D003198:Computer Simulation; D017124:Conserved Sequence; D051048:Diacylglycerol O-Acyltransferase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019143:Evolution, Molecular; D005288:Ferredoxins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D053858:Metabolic Networks and Pathways / Q000941:ethics; D010802:Phylogeny; D056510:Position-Specific Scoring Matrices; D014280:Triglycerides / Q000096:biosynthesis / Q000378:metabolism*
chemical_list: D005288:Ferredoxins; D014280:Triglycerides; D051048:Diacylglycerol O-Acyltransferase
keywords: Algae; Biodiesel production; Chloroplast; HMMER profiling; Neutral lipids; Protein phylogeny; Soluble acyltransferase; Triglyceride metabolism