Gene structure and M20T polymorphism of the Schistosoma mansoni Sm14 fatty acid-binding protein. Molecular, functioanl, and immunoprotection analysis.
The Schistosoma mansoni Sm14 antigen belongs to the fatty acid-binding protein family and is considered a vaccine candidate against at least two parasite worms, Fasciola hepatica and S. mansoni. Here the genomic sequence and the polymorphism of Sm14 have been characterized for the first time. We found that the conserved methionine at position 20 is polymorphic, being exchangeable with threonine (M20T). To evaluate the function of the amino acid residue at this position, we have also constructed the mutant Sm14-A20 besides the two native isoforms (Sm14-M20 and Sm14-T20). The three purified recombinant His(6)-tagged Sm14 proteins (rSm14-M20, rSm14-T20, and rSm14-A20) present a predominant beta-barrel structure as shown by CD spectroscopy. Thermal and urea unfolding studies evidenced a higher structural stability of rSm14-M20 over the other forms (rSm14-M20>rSm14-T20>rSm14-A20). All of the Sm14 proteins were able to bind 11-(dansylamino)undecanoic acid (DAUDA) without substantial difference in the binding affinity. However, rSm14-M20 exhibited a higher affinity for natural fatty acids than the rSm14-T20 and rSm14-A20 proteins as judged by competitive experiments against DAUDA (rSm14-M20>rSm14-T20>rSm14-A20). The rSm14-M20 or rSm14-T20 isoforms but not the rSm14-A20 mutant was able to induce significant protection against S. mansoni cercariae challenge in immunized mice. The level of protection efficacy correlates with the extent of structure stability of the recombinant Sm14 isoforms and mutant.
Ramos
Celso Raul Romero
CR
Instituto Butantan, Avenida Vital Brasil 1500, 05503-900, São Paulo, Brazil.
Figueredo
Rita Cassia Rossi
RC
Pertinhez
Thelma Aguiar
TA
Vilar
Mônica Magno
MM
do Nascimento
Ana Lúcia Tabet Oller
AL
Tendler
Miriam
M
Raw
Isaías
I
Spisni
Alberto
A
Ho
Paulo Lee
PL
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001483:Base Sequence; D001938:Brazil; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D002942:Circular Dichroism; D017931:DNA Primers; D050612:Fatty Acid Transport Proteins; D050556:Fatty Acid-Binding Proteins; D005260:Female; D005843:Geography; D015801:Helminth Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D007438:Introns; D008297:Male; D026901:Membrane Transport Proteins; D008969:Molecular Sequence Data; D005810:Multigene Family; D020125:Mutation, Missense; D009363:Neoplasm Proteins; D011110:Polymorphism, Genetic; D012550:Schistosoma mansoni / Q000235:genetics*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013816:Thermodynamics
D002352:Carrier Proteins; D017931:DNA Primers; D050612:Fatty Acid Transport Proteins; D050556:Fatty Acid-Binding Proteins; D015801:Helminth Proteins; D026901:Membrane Transport Proteins; D009363:Neoplasm Proteins; C068709:SM14 protein, Schistosoma mansoni