Purification of the paraflagellar rod of the trypanosomatid Herpetomonas megaseliae and identification of some of its minor components.
The paraflagellar rod (PFR) is a component of the flagellar cytoskeleton of trypanosomatid protozoa, representing a filamentous structure that runs alongside the common 9 + 2 microtubular axoneme. The high degree of ultrastructural complexity and organization of the PFR suggests that it might be formed by numerous biochemical components. However, biochemical analysis of the PFR has revealed, to date, a modest degree of complexity in what concerns both major and minor PFR proteins. In this paper the preparation of purified PFR fractions by a combination of conventional cell-fractionation procedures, non-ionic detergent treatment and limited proteolysis is described. Comparative SDS-PAGE analysis of the different purification steps indicates that the purified PFR fractions possess high amounts of the well-known major PFR proteins (77 and 83 kDa). Also, bands of 147, 139, 129 and 122 kDa are clearly enriched in such fractions and may correspond to minor PFR components. A slight enrichment in a specific fraction of a doublet of bands of 181/188 kDa suggest the participation of these proteins in the composition of the bridges between the PFR and the axoneme.
Moreira-Leite
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Instituto de Biofisica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, RJ, Brazil.
de Souza
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da Cunha-e-Silva
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D000818:Animals; D002458:Cell Fractionation; D003599:Cytoskeleton / Q000648:ultrastructure*; D005407:Flagella / Q000648:ultrastructure*; D013347:Subcellular Fractions / Q000648:ultrastructure; D014351:Trypanosomatina / Q000648:ultrastructure*