Nitrite-oxidizing chemoautotrophic Nitrobacter agilis ATCC 14123 exhibited remarkedly strong ribulose-1,5-bisphosphate carboxylase (RuBisCO) activity. RuBisCO [EC 4.1.1.39] was purified as an electrophoretically homogeneous protein. The molecular mass of the enzyme was estimated to be about 480 kDa by gel filtration, suggesting that the enzyme consisted of two different subunits [Large (L) : 50 kDa, Small (S) : 10 kDa], as demonstrated by SDS-PAGE. This confirmed that the enzyme has a L(8)S(8) structure. The K-m values of the enzyme for RuBP, NaHCO3, and Mg2+ were estimated to be 0.039 mM, 6.39 mM and 1.36 mM, respectively. The optimum pH and temperature for its activity were approximately 8.0 and 40 degrees C, respectively. The enzyme was stable up to 45 degrees C and in a pH range of 7.0-9.0 (4 degrees C, 48 h). The enzyme activity was inhibited by Mo2+, CU2+, Hg2+, N-ethylmaleimide, p-chloromercuribenzoate and SDS (1 mM). The N-terminal amino acid sequence of the large subunit is AVKSYQAGVTQYRQSYWQPDYMPL, and that of the small subunit is AVQAYRSLKKYETFSYLPQ.