SMase II, a new sphingomyelinase D from Loxosceles laeta venom gland: molecular cloning, expression, function and structural analysis.
Sphingomyelinase D (SMase D) present in the venoms of Loxosceles spiders is the principal component responsible for local and systemic effects observed in the loxoscelism. By using "expressed sequencing tag", it was possible to identify, in a L. laeta venom gland library, clones containing inserts coding for proteins with similarity to SMase D. One of these clones was expressed and the recombinant protein compared with the previously characterized SMase I from L. laeta, in terms of their biological, biochemical and structural properties. The new recombinant protein, SMase II, possesses all the biological properties ascribed to the whole venom and SMase I. SMase II shares 40% and 77% sequence similarity with SMase I and Lb3, respectively; the latter, a SMase D isoform from L. boneti, catalytically inactive. Molecular modeling and molecular dynamics simulations were employed to understand the structural basis, especially the presence of an additional disulfide bridge, in an attempt to account for the observed differences in SMases D activity.
de Santi Ferrara
Guilherme I
GI
Laboratório de Imunoquímica, Instituto Butantan, São Paulo, Brazil.
Fernandes-Pedrosa
Matheus de F
Mde F
Junqueira-de-Azevedo
Inácio de L M
Ide L
Gonçalves-de-Andrade
Rute M
RM
Portaro
Fernanda C V
FC
Manzoni-de-Almeida
Daniel
D
Murakami
Mário T
MT
Arni
Raghuvir K
RK
van den Berg
Carmen W
CW
Ho
Paulo L
PL
Tambourgi
Denise V
DV
D000595:Amino Acid Sequence; D015153:Blotting, Western; D002021:Buffers; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004912:Erythrocytes / Q000187:drug effects; D005088:Exocrine Glands / Q000201:enzymology*; D005434:Flow Cytometry; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D007202:Indicators and Reagents; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009336:Necrosis / Q000139:chemically induced / Q000473:pathology; D010727:Phosphoric Diester Hydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry; D012867:Skin / Q000473:pathology; D013111:Spider Venoms / Q000201:enzymology*
D002021:Buffers; D007202:Indicators and Reagents; D011994:Recombinant Proteins; D013111:Spider Venoms; D010727:Phosphoric Diester Hydrolases; C055160:sphingomyelin phosphodiesterase D