Candidates of trichocyst matrix proteins of the dinoflagellate Oxyrrhis marina.

Trichocysts are a common cell organelle of ciliates and dinoflagellates. They are composed of trichocyst matrix proteins and have been intensely investigated and characterized in ciliates. Here, for the first time, data have been obtained for trichocyst matrix proteins of a dinoflagellate. A DELTA-BLAST search using 14 available and complete amino acid sequences of mature trichocyst matrix proteins of the ciliate Paramecium tetraurelia resulted in 16 hits for the dinoflagellate Oxyrrhis marina when the E values and bit values to be scored were <10 and >40. They code for proteins with acidic pI values and exceeded the precursors of the trichocyst matrix proteins of the ciliate approximately twofold in length. The values calculated for coverage, identity, and positives ranged from 76 to 100, 21.5 to 28.3, and 44.9 to 53.9%, respectively. Protein conformation predictions indicate coiled-coil domains which are a common feature of mature ciliate trichocyst matrix proteins. As often several EST sequences of O. marina matched with a queried mature trichocyst matrix protein of P. tetraurelia, a multigene family can be assumed for trichocyst proteins in this dinophyte, too. Trichocyst-enriched fractions of O. marina were isolated and subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis. When samples were incubated with loading buffer without a reducing agent, the banding pattern was mainly composed of three regions in the range of >90, 75-60, and 50-35 kDa, with each region consisting of four to five bands. Tryptic in gel digestion of proteins excised from these three gel regions followed by mass spectrometry confirmed that up to 14 of the 16 predicted proteins were present within the trichocyst-enriched fractions. When the samples were reduced with either ß-mercaptoethanol or dithiothreitol, the proteins of the three regions disappeared almost completely and proteins in the range of 27 to 15 kDa became the dominating bands. Up to 12 of the predicted proteins were detected within these bands.