title: Molecular cloning of a hyaluronidase from Bothrops pauloensis venom gland.
abstract: BACKGROUND
  Hyaluronate is one of the major components of extracellular matrix from vertebrates whose breakdown is catalyzed by the enzyme hyaluronidase. These enzymes are widely described in snake venoms, in which they facilitate the spreading of the main toxins in the victim's body during the envenoming. Snake venoms also present some variants (hyaluronidases-like substances) that are probably originated by alternative splicing, even though their relevance in envenomation is still under investigation. Hyaluronidases-like proteins have not yet been purified from any snake venom, but the cDNA that encodes these toxins was already identified in snake venom glands by transcriptomic analysis. Herein, we report the cloning and in silico analysis of the first hyaluronidase-like proteins from a Brazilian snake venom.
  
  
  METHODS
  The cDNA sequence of hyaluronidase was cloned from the transcriptome of Bothrops pauloensis venom glands. This sequence was submitted to multiple alignment with other related sequences by ClustalW. A phylogenetic analysis was performed using MEGA 4 software by the neighbor joining (NJ) method.
  
  
  RESULTS
  The cDNA from Bothrops pauloensis venom gland that corresponds to hyaluronidase comprises 1175 bp and codifies a protein containing 194 amino acid residues. The sequence, denominated BpHyase, was identified as hyaluronidase-like since it shows high sequence identities (above 83%) with other described snake venom hyaluronidase-like sequences. Hyaluronidases-like proteins are thought to be products of alternative splicing implicated in deletions of central amino acids, including the catalytic residues. Structure-based sequence alignment of BpHyase to human hyaluronidase hHyal-1 demonstrates a loss of some key secondary structures. The phylogenetic analysis indicates an independent evolution of BpHyal when compared to other hyaluronidases. However, these toxins might share a common ancestor, thus suggesting a broad hyaluronidase-like distribution among venomous snakes.
  
  
  CONCLUSIONS
  This work is the first report of a cDNA sequence of hyaluronidase from Brazilian snake venoms. Moreover, the in silico analysis of its deduced amino acid sequence opens new perspectives about the biological function of hyaluronidases-like proteins and may direct further studies comprising their isolation and/or recombinant production, as well as their structural and functional characterization.
authors.affiliation: Instituto de Genética e Bioquímica, Universidade Federal de Uberlândia, Uberlândia, MG CEP 384000-902, Brasil ; National Institute of Sciences and Technology on Nanobiopharmaceutics (INCT), Federal University of Minas Gerais (UFMG), Belo Horizonte, Minas Gerais State, Brazil.
authors.forename: Letícia Eulalio
authors.initials: LE
authors.lastname: Castanheira
authors.affiliation: Instituto de Genética e Bioquímica, Universidade Federal de Uberlândia, Uberlândia, MG CEP 384000-902, Brasil ; National Institute of Sciences and Technology on Nanobiopharmaceutics (INCT), Federal University of Minas Gerais (UFMG), Belo Horizonte, Minas Gerais State, Brazil.
authors.forename: Renata Santos
authors.initials: RS
authors.lastname: Rodrigues
authors.affiliation: Department of Physics and Chemistry, School of Pharmaceutical Sciences, University of São Paulo (USP), Ribeirão Preto, São Paulo State, Brazil.
authors.forename: Johara
authors.initials: J
authors.lastname: Boldrini-França
authors.affiliation: Department of Genetics and Evolution, Federal University of São Carlos (UFSCar), São Carlos, São Paulo State, Brazil.
authors.forename: Fernando Pp
authors.initials: FP
authors.lastname: Fonseca
authors.affiliation: Department of Genetics and Evolution, Federal University of São Carlos (UFSCar), São Carlos, São Paulo State, Brazil.
authors.forename: Flávio
authors.initials: F
authors.lastname: Henrique-Silva
authors.affiliation: Instituto de Genética e Bioquímica, Universidade Federal de Uberlândia, Uberlândia, MG CEP 384000-902, Brasil.
authors.forename: Maria I
authors.initials: MI
authors.lastname: Homsi-Brandeburgo
authors.affiliation: Instituto de Genética e Bioquímica, Universidade Federal de Uberlândia, Uberlândia, MG CEP 384000-902, Brasil ; National Institute of Sciences and Technology on Nanobiopharmaceutics (INCT), Federal University of Minas Gerais (UFMG), Belo Horizonte, Minas Gerais State, Brazil.
authors.forename: Veridiana M
authors.initials: VM
authors.lastname: Rodrigues
keywords: Alternative splicing; Hyaluronidase-like; Snake venom