Cloning, sequencing and functional expression of cytosolic malate dehydrogenase from Taenia solium: Purification and characterization of the recombinant enzyme

[Display omitted] ► The complete coding sequence of a Taenia solium malate dehydrogenase is reported. ► The gene is composed of two 498pb exons, and one 76pb intron. ► It was produced a T. solium malate dehydrogenase recombinant protein. ► A partial characterization of the recombinant protein is rep...

Full description

Saved in:
Bibliographic Details
Published in:Experimental parasitology 2011-07, Vol.128 (3), p.217-224
Main Authors: Nava, Gabriela, Laclette, Juan P., Bobes, Raúl, Carrero, Julio C., Reyes-Vivas, Horacio, Enriquez-Flores, Sergio, Mendoza-Hernández, Guillermo, Plancarte, Agustín
Format: Article
Language:English
Subjects:
Amino Acid Sequence
amino acid sequences
amino acids
Animals
Base Sequence
Biological and medical sciences
Circular Dichroism
Cloning
Cloning, Molecular
complementary DNA
Cysticercosis
Cytosol - enzymology
DNA, Complementary - chemistry
enzyme activity
Enzyme Stability
Fundamental and applied biological sciences. Psychology
gel electrophoresis
Gene Expression Regulation, Enzymologic
genes
Hydrogen-Ion Concentration
Isoelectric Focusing
isoelectric point
Kinetics
lactate dehydrogenase
Life cycle. Host-agent relationship. Pathogenesis
Malate dehydrogenase
Malate Dehydrogenase - chemistry
Malate Dehydrogenase - genetics
Malate Dehydrogenase - isolation & purification
Malate Dehydrogenase - metabolism
malates
Metabolism
Molecular Sequence Data
molecular weight
NAD (coenzyme)
NAD - metabolism
NADP (coenzyme)
oxidation
parasitology
Protozoa
Recombinant enzyme
recombinant proteins
Recombinant Proteins - chemistry
Recombinant Proteins - genetics
Recombinant Proteins - isolation & purification
Recombinant Proteins - metabolism
Sequence Alignment
Sequence Analysis
Taenia solium
Taenia solium - enzymology
Taenia solium - genetics
Taeniasolium
Temperature
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:[Display omitted] ► The complete coding sequence of a Taenia solium malate dehydrogenase is reported. ► The gene is composed of two 498pb exons, and one 76pb intron. ► It was produced a T. solium malate dehydrogenase recombinant protein. ► A partial characterization of the recombinant protein is reported. We report herein the complete coding sequence of a Taenia solium cytosolic malate dehydrogenase (TscMDH). The cDNA fragment, identified from the T. solium genome project database, encodes a protein of 332 amino acid residues with an estimated molecular weight of 36517Da. For recombinant expression, the full length coding sequence was cloned into pET23a. After successful expression and enzyme purification, isoelectrofocusing gel electrophoresis allowed to confirm the calculated pI value at 8.1, as deduced from the amino acid sequence. The recombinant protein (r-TscMDH) showed MDH activity of 409U/mg in the reduction of oxaloacetate, with neither lactate dehydrogenase activity nor NADPH selectivity. Optimum pH for enzyme activity was 7.6 for oxaloacetate reduction and 9.6 for malate oxidation. Kcat values for oxaloacetate, malate, NAD, and NADH were 665, 47, 385, and 962s−1, respectively. Additionally, a partial characterization of TsMDH gene structure after analysis of a 1.56Kb genomic contig assembly is also reported.
ISSN:0014-4894
1090-2449
DOI:10.1016/j.exppara.2011.03.008