Molecular cloning, expression and immunological properties of LiD1, a protein from the dermonecrotic family of Loxosceles intermedia spider venom.
The present report describes the identification and molecular characterization of LiD1, a protein expressed in the venom gland of the brown spider Loxosceles intermedia. LiD1 belongs to a family of proteins with dermonecrotic activity and members of this family have been found in spiders from the genus Loxosceles. The necrotic lesions caused by this group of proteins may lead to serious socio-economic problems such as surgical tissue reconstitution and even patient death. LiD1 was cloned using a cDNA library constructed from the venom gland of L. intermedia and antibodies against proteins with dermonecrotic activity isolated from the crude venom of this spider. The amino acid sequence deduced from the cDNA revealed a mature protein of approximately 31 kDa, with a pI of 7.37. The cDNA also revealed the existence of a signal peptide, a propeptide and also an untranslated 3' region with 218 nucleotides. LiD1 was expressed as a protein fused with beta-galactoside protein using the vector pBK-CMV, resulting in the recombinant protein recLiD1 with important immunological properties. recLiD1 was strongly recognised by anti-dermonecrotic antibodies and was also able to generate reactive antibodies against native dermonecrotic proteins isolated from the venom of L. intermedia.
Kalapothakis
Evanguedes
E
Departamento de Farmacologia, Divisao de Biologia Celular, Universidade Federal de Minas Gerais, Av. Antonio Carlos 6627, Belo Horizonte, MG, Cep 31270901, Brazil. ekalapo@icb.ufmg.br
Araujo
Simone Costa
SC
de Castro
Cibele Soares
CS
Mendes
Thais Melo
TM
Gomez
Marcus Vinícius
MV
Mangili
Oldemir C
OC
Gubert
Ida C
IC
Chávez-Olórtegui
Carlos
C
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D005591:Chemical Fractionation; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D015870:Gene Expression; D015723:Gene Library; D008969:Molecular Sequence Data; D010727:Phosphoric Diester Hydrolases / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics; D013111:Spider Venoms / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D013112:Spiders / Q000502:physiology*
D018076:DNA, Complementary; D011994:Recombinant Proteins; D013111:Spider Venoms; C055161:loxosceles venom; D010727:Phosphoric Diester Hydrolases