Figures & data
Figure 1. The CO2 hydration activity of SFHb and PolySFHb was assayed. Six concentrations between 0.2–10.5 mg/mL of sample were selected in order to obtain results in an appropriate and measurable range. The CA activity was described in W–A units. Measurements were done in triplicates to indicate statistical reproducibility.
Figure 2. (A) PolySFHb was prepared from SFHb and crosslinked with increasing amount of CA (200 units/mL and 400 units/mL). Enzyme activity of SFHb and PolySFHb prepared from SFHb without additional CA was also assayed. (B) PolySFHb was prepared from SFHb and was assayed for CA activity. The CO2 hydration activity of PolySFHb crosslinked with CA (1070 units/mL) was also measured.
Figure 3. Samples containing CA in free form and samples without free enzyme were assayed for CA activity. The expected activity for the mixture of PolySFHb and free form CA is also indicated.
Figure 4. PolySFHb was mixed with CA in free form and assayed for enzyme activity. PolySFHb without additional enzyme and CA alone were also assayed. The sample concentrations were proportionally reduced and selected to be between 0–0.35 mg/mL.
Figure 5. PolySFHb was prepared from SFHb and assayed with the electrometric delta pH assay using different concentrations of bicarbonate and carbonate buffers corresponding to physiological conditions.
Figure 6. (A) Plot illustrating the Michaelis-Menten equation for CA in free form. The curve was obtained by increasing the substrate concentration until a stable rate of CO2 hydration was obtained. (B) Lineweaver-Burk plot for CA in free form. The curve was obtained by taking the reciprocal of the substrate and the rate of enzyme activity.
Figure 7. (A) Plot illustrating the Michaelis-Menten equation for SFHb. (B) Lineweaver-Burk plot for SFHb.
Figure 8. (A) Plot illustrating the Michaelis-Menten equation for PolySFHb-CA. (B) Lineweaver-Burk plot for PolySFHb-CA.
Table 1. Apparent kinetic constants for CA in free form, SFHb, and PolySFHb-CA.
Figure 9. Superoxide dismutase activity of SFHb, PolySFHb, SFHb+SOD+CAT+CA and polySFHb-SOD-CAT-CA. SOD (1050 units/mL), catalase (21,000 units/mL), and carbonic anhydrase (1070 units/mL) were added to stroma-free hemoglobin (7 g/dl), then polymerized into PolySFHb-SOD-CAT-CA, resulting in an Hb: SOD ratio of 1g: 18,000 after crosslinking.
Figure 10. catalase activity of SFHb, PolySFHb, SFHb+SOD+CAT+CA and polySFHb-SOD-CAT-CA. SOD (1050 units/mL), catalase (21,000 units/mL), and carbonic anhydrase (1070 units/mL) were added to stroma-free hemoglobin (7 g/dl), then polymerized into PolySFHb-SOD-CAT-CAT, resulting in an Hb:CAT ratio of 1g: 130,000U after crosslinking.
Figure 11. Carbonic anhydrase activity of SFHb, PolySFHb, SFHb+SOD+CAT+CA and polySFHb-SOD-CAT-CA. SOD (1050 units/mL), catalase (21,000 units/mL), and carbonic anhydrase (1070 units/mL) were added to stroma-free hemoglobin (7 g/dl), then polymerized into PolySFHb-SOD-CAT-CA, resulting in an Hb:CA ratio of 1g: 130,000U after crosslinking.
Table 2. Molecular distribution of PolySFHb-SOD-CAT-CA and enzyme activities.
