The proteins of the 30 S ribosomal subunits from spinach chloroplasts were investigated using a radical-free and highly reducing (RFHR) method of two-dimensional polyacrylamide gel electrophoresis (PAGE). Twenty-three proteins were resolved on the gel down to the smallest protein of 5 kDa. The N-terminal amino acid sequence of the 5 kDa protein showed no homology with that of any other protein stored in databases, and the copy numbers were estimated to be 0.88 +/- 0.16 and 0.72 +/- 0.04 in the 30 S subunits and the 70 S ribosomes, respectively. The results suggest that the 5 kDa protein, which we have called SCS23, may be an essential ribosomal protein specific to spinach chloroplasts.