Carbonic anhydrase purified from the cell surface of Chlamydomonas reinhurdtii was inactivated by treatment with dithiothreitol. This treatment caused dissociation of the holoenzyme into 35-kDa (A) and 4-kDa (B) subunits as revealed by SDS/PAGE. The 35-kDa subunit was further separated into two components Al (35 kDa) and A2 (36.5 kDa) by SDSjPAGE using a gradient gel. These two components have the same amino acid sequence up to at least the 10th amino acid from the N-terminus. The molecular masses were estimated at 76 kDa and 35 kDa for the holoenzyme and the large subunit, respectively, and the molar ratio of the former to the latter at 1 : 2, by using the techniques of low-angle laser light-scattering photometry and precision differential refractometry combined with gel-filtration HPLC. The molar ratio of the 35-kDa/4-kDa subunits was estimated at 1 : l the gelfiltration HPLC monitored with precision differential refractometry. Atomic-absorption spectrophotometry revealed that the holoenzyme contains two atoms of zinc. These results suggest that the holoenzyme is a heterotetramer composed of two large subunits (A, and A,) and two small subunits (B).Carbonic anhydrase catalyzes the reversible hydration of CO, dissolved in water. The activity of carbonic anhydrase in the cells of Chlumydomonas reinhardtii increases during adaptation from C0,-enriched air to ordinary air [I, 21. The same phenomenon has been reported in various species of microalgae (for a review see [3]). In Chlamydomonas cells, carbonic anhydrase is synthesized de novo during adaptation (4, 51. A 42-kDa precursor polypeptide is first synthesized and its molecular mass is quickly decreased to 35 kDa in the cell [6]. Most of the enzyme activity is located outside the plasma membrane and can be released into the suspending medium by treating the cells with a gametic wall-lysing enzyme [7] or trypsin [S]. It is easily solubilized when cells are broken, and has been purified as a glycoprotein [6, 91. Based on the results obtained with SDSjPAGE and gel-filtration HPLC, it was previously assumed that carbonic anhydrase of Chlamydomonas cells was composed of three identical subunits with a molecular mass of 35 kDa [lo]. In the present paper, however, we report the presence of a 4-kDa subunit in addition to the 35-kDa subunit in Chlamydomonas carbonic anhydrase, and propose a heterologous subunit constitution.

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Amorphous ceramics with the composition of Bi2Pb0.4Sr2Ca2Cu3O y were prepared by rapid quenching. The change in crystalline structure during annealing and the effect of the “seeding” with the high-T c phase were investigated. The volume fraction of the high-T c phase increases with annealing time; however, the low-T c phase reaches a maximum value of 38% for 1 h annealing and decreases on prolonged annealing. The formation rate of the high-T c phase changes when the volume of the high-T c phase reaches about 30%. TEM observation shows that the high-T c phase is often located between the low-T c phase and the nonsuperconducting phase. The addition of the seed crystals with the high-T c phase is effective in forming the high-T c phase in shorter annealing times, but a smaller volume fraction of the high-T c phase was obtained after annealing for 50 h.

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