INFLUENCE OF THE A-HELIX STRUCTURE ON THE POLYMERIZATION OF HEMOGLOBIN-S

Hb S variants containing Lys-beta 132 --> Ala or Asn substitutions wer e engineered to evaluate the consequences of the A helix destabilizati on in the polymerization process. Previous studies suggested that the loss of the Glu-beta 7-Lys-beta 132 salt bridge in the recombinant Hb beta E6V/E7A could be responsible for the destabilization of the A hel ix. The recombinant Hb (rHb) S/beta 132 variants polymerized with an i ncreased delay time as well as decreased maximum absorbance and Hb sol ubility values similar to that of Hb S. These data indicate that the s trength of the donor-acceptor site interaction may be reduced due to a n altered conformation of the A helix. The question arises whether thi s alternation leads to a true inhibition of the polymerization process or to qualitatively different polymers. The oxygen affinity of the be ta 132 mutated rHbs was similar to that of Hb A and S, whereas the coo peratively and effects of organic phosphates were reduced. This could be attributed to the modifications in the central cavity due to loss o f the positively charged lysine. Since Lys-beta 132 is involved in the stabilization of the alpha 1-beta 1 interface, the loss of the beta 1 32(H10)-beta 128(H6) salt bridge may be responsible for the marked the rmal instability of the beta 132 mutated rHbs.