2-DIMENSIONAL GEL-ELECTROPHORESIS OF CAENORHABDITIS-ELEGANS HOMOGENATES AND IDENTIFICATION OF PROTEIN SPOTS BY MICROSEQUENCING

Employing isoelectric focusing on immobilized pH gradients followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) we have obtained a map of C. elegans proteins, from a mixed culture co ntaining all developmental stages, presenting over 2000 spots within t he window of isoelectric points (pr) 3.5-9 and a molecular mass of 10- 200 kDa. Edman microsequencing yielded successful results in 12 out of 24 analyzed spots. All but one of the N-terminal sequences retrieved C. elegans sequences in cosmid and/or expressed sequence tag clones. S tructurally related protein sequences found in data banks included enz ymes in energy metabolism (cytochrome oxydase, ATP synthase, enolase), a fatty acid-binding protein, a translationally controlled tumor prot ein, an unknown C. elegans protein, an acidic ribosomal protein, a tit in-like protein, a G-protein beta chain, cyclophilin, and cathepsin D. Experimental determination of N-termini allowed us to define sites of signal cleavage providing further information on the physiological ro le of the newly found C. elegans proteins. This report demonstrates th e possibility of two-dimensional gel electrophoresis and Edman microse quencing in the elucidation of C. elegans proteome.