Production of bioactive conjugated linoleic acid by the multifunctional enolase from Lactobacillus plantarum.
Lactobacillus plantarum α-enolase, a multifunctional-anchorless-surface protein belonging to the conserved family of enolases with a central role in glycolytic metabolism, was characterized to have a side role in the intricate metabolism of biohydrogenation of linoleic acid, catalyzing the formation of bioactive 9-cis-11-trans-CLA through dehydration and isomerization of 10-hydroxy-12-cis-octadecenoic acid. The identity of the enolase was confirmed through mass spectrometric analysis that showed the characteristic 442 amino acid sequence with a molecular mass of 48.03kDa. The enolase was not capable of using linoleic acid directly as a substrate but instead uses its hydroxyl derivative 10-hydroxi-12-cis-octadecenoic acid to finally form bioactive conjugated linoleic acid. Biochemical optimization studies were carried out to elucidate the conditions for maximum production of 9-cis-11-trans-CLA and maximum stability of α-enolase when catalyzing this reaction. Furthermore, through structural analysis of the protein, we propose the binding sites of substrate and product molecules that were characterized as two hydrophobic superficial pockets located at opposite ends of the enolase connected through a channel where the catalysis of dehydration and isomerization might occur. These results prove that multifunctional α-enolase also plays a role in cell detoxification from polyunsaturated fatty acids such as linoleic acid, along with the linoleate isomerase complex.
Departamento de Química de Biomacromoléculas, Instituto de Química, Universidad Nacional Autónoma de México, Circuito Exterior, Ciudad Universitaria, Coyoacán, C.P. 04510, México. Electronic address: joanna@comunidad.unam.mx.
Joana
J
Ortega-Anaya
Departamento de Química de Biomacromoléculas, Instituto de Química, Universidad Nacional Autónoma de México, Circuito Exterior, Ciudad Universitaria, Coyoacán, C.P. 04510, México. Electronic address: hersan@unam.mx.
Alejandra
A
Hernández-Santoyo
D000595:Amino Acid Sequence; D001665:Binding Sites; D002849:Chromatography, Gas; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007536:Isomerism; D048191:Lactiplantibacillus plantarum / Q000201:enzymology*; D044243:Linoleic Acids, Conjugated / Q000096:biosynthesis*; D008958:Models, Molecular; D009829:Oleic Acids / Q000737:chemistry / Q000378:metabolism; D010751:Phosphopyruvate Hydratase / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D000072417:Protein Domains; D020539:Sequence Analysis, Protein; D013379:Substrate Specificity; D013696:Temperature
D044243:Linoleic Acids, Conjugated; D009829:Oleic Acids; C040430:10-hydroxy-12-octadecenoic acid; D010751:Phosphopyruvate Hydratase
10-Hydroxy-12-cis-octadecenoic acid; Conjugated linoleic acid; Dehydration; Enolase; Isomerization; Lactobacillus plantarum