Characterization of a blood activated chitinolytic system in the midgut of the sand fly vectors Lutzomyia longipalpis and Phlebotomus papatasi

J. M. Ramalho-Ortigão, S. Kamhawi, M. B. Joshi, David Reynoso, P. G. Lawyer, D. M. Dwyer, D. L. Sacks, J. G. Valenzuela

Research output: Contribution to journalArticle

33 Citations (Scopus)

Abstract

We characterized a cDNA from Phlebotomus papatasi, PpChit1, which encodes a midgut specific chitinase and show the presence of a functional, blood-induced chitinolytic system in sand flies. PpChit1 is detected only in the midgut and is regulated by blood feeding. A recombinant protein (rPpChit1) produced in HEK 293-F cells exhibited a similar activity profile to that found in the native protein against several specific substrates, including an oligomeric glycol chitin and synthetic 4-methyl-umbelliferone labelled substrates. Western blotting showed that the native protein is recognized by mouse polyclonal antibodies against rPpChit1. Additionally, the rPpChit1 and the native chitinase displayed similar retention times in a HPLC size fractionation column. When added to rPpChit1 or to midgut lysates, PpChit1 sera reduced chitinolytic activity by 65-70%.

Original languageEnglish (US)
Pages (from-to)703-712
Number of pages10
JournalInsect Molecular Biology
Volume14
Issue number6
DOIs
StatePublished - Dec 1 2005
Externally publishedYes

Fingerprint

Phlebotomus
Phlebotomus papatasi
Lutzomyia longipalpis
Psychodidae
Chitinases
7-hydroxycoumarin
Phlebotominae
midgut
chitinase
HEK293 Cells
blood
Recombinant Proteins
glycols
Complementary DNA
Western Blotting
High Pressure Liquid Chromatography
chitin
blood serum
polyclonal antibodies
recombinant proteins

Keywords

  • Chitinolytic activity
  • Leishmania
  • Peritrophic matrix
  • Sand fly

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics
  • Insect Science

Cite this

Characterization of a blood activated chitinolytic system in the midgut of the sand fly vectors Lutzomyia longipalpis and Phlebotomus papatasi. / Ramalho-Ortigão, J. M.; Kamhawi, S.; Joshi, M. B.; Reynoso, David; Lawyer, P. G.; Dwyer, D. M.; Sacks, D. L.; Valenzuela, J. G.

In: Insect Molecular Biology, Vol. 14, No. 6, 01.12.2005, p. 703-712.

Research output: Contribution to journalArticle

Ramalho-Ortigão, J. M. ; Kamhawi, S. ; Joshi, M. B. ; Reynoso, David ; Lawyer, P. G. ; Dwyer, D. M. ; Sacks, D. L. ; Valenzuela, J. G. / Characterization of a blood activated chitinolytic system in the midgut of the sand fly vectors Lutzomyia longipalpis and Phlebotomus papatasi. In: Insect Molecular Biology. 2005 ; Vol. 14, No. 6. pp. 703-712.
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