Cloning and expression of chitinases of Entamoebae

Humberto De La Vega, Charles A. Specht, Carlos E. Semino, Phillips W. Robbins, Daniel Eichinger, Daniel Caplivski, Sudip Ghosh, John Samuelson

Research output: Contribution to journalArticlepeer-review

67 Scopus citations

Abstract

Entamoeba histolytica (Eh) and Entamoeba dispar (Ed) are protozoan parasites that infect hundreds of millions of persons. In the colonic lumen, amebae form chitin-walled cysts, the infectious stage of the parasite. Entamoeba ivadens (Ei), which infects reptiles and is a model for amebic encystation, produces chitin synthase and chitinase during encystation. Ei cyst formation is blocked by the chitinase-inhibitor allosamidin. Here molecular cloning techniques were used to identify homologous genes of Eh, Ed, and Ei that encode chitinases (EC 3.2.1.14). The Eh gene (Eh cht 1) predicts a 507-amino acid (aa) enzyme, which has 93 and 74% positional identities with Ed and Ei chitinases, respectively. The Entamoeba chitinases have signal sequences, followed by acidic and hydrophilic sequences composed of multiple tandemly arranged 7-aa repeats (Eh and Ed) or repeats varying in length (Ei). The aa compositions of the chitinase repeats are similar to those of the repeats of the Eh and Ed Ser-rich proteins. The COOH-terminus of each chitinase has a catalytic domain, which resembles those of Brugia malayi (33% positional identity) and Manduca sexta (29%). Recombinant Entamoeba chitinases are precipitated by chitin and show chitinase activity with chitooligosacharide substrates. Consistent with previous biochemical data, chitinase mRNAs are absent in Ei trophozoites and accumulate to maximal levels in Ei encysting for 48 h.

Original languageEnglish
Pages (from-to)139-147
Number of pages9
JournalMolecular and Biochemical Parasitology
Volume85
Issue number2
DOIs
StatePublished - Apr 1997
Externally publishedYes

Keywords

  • Entamoeba dispar
  • Entamoeba histolytica
  • Entamoeba invadens
  • chitinase
  • cysts
  • hydrophilic repeats

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