SUMMARY: ThiobacillusA2 grew on formate as its sole source of carbon and energy. Growth was autotrophic: formate and carbon dioxide were equivalent as carbon sources during growth on formate. Carbon dioxide was fixed by the Calvin cycle in formate- or thiosulphate-grown bacteria which contained comparable high specific activities of ribulosebisphosphate carboxylase. Formate incorporated by bacteria growing heterotrophically or on thio- sulphate showed more restricted metabolism, particularly providing carbon for purines. Bacteria growing on formate or thiosulphate assimilated acetate but showed disproportionately high incorporation into glutamate, proline, arginine, leucine, pyrimidines and lipid. Growth kinetics on formate were studied using extended cultures held at constant formate concentrations at pH 7.8. Yield was a function of substrate concentration and growth rate, which were linearly related in the range 4 to 40 mM-formate. Formate was an inhibitory substrate at higher concentrations. A computer analysis of the inhibited growth kinetics indicated Ks and Ki values of 26.5 and 187 mM-formate, respectively, and a true μ max of 0.299 h−1.