Inhibition of the Biosynthetic Completion of Rabbit Hemoglobin by Isolated Human Hemoglobin Chains

Abstract

Abstract A cell-free system from rabbit reticulocytes incorporated 14C-valine and 14C-leucine into hemoglobin. This synthesis of rabbit hemoglobin was inhibited in a specific manner when α or β chains, isolated from human hemoglobin, were added to the system. The human α and β chains interacted with the newly synthesized 14C-rabbit chains to form hybrid hemoglobins, which were identified by cellulose acetate electrophoresis, and the amount of 14C-rabbit hemoglobin was decreased. When human β chains were added to the reaction, there was also a 40 to 50% decrease in the amount of the 14C-rabbit β chain component in the supernatant fraction and an increase in the amount of completed 14C-β chains in the ribosome fraction, as determined by chain separation chromatography of these fractions on carboxymethyl cellulose columns. The supernatant phase also contained an electrophoretic fraction with the properties of free 14C-rabbit β chains; this fraction was not detected in the system incubated without human β chains. Addition of hemin at the beginning of the reaction largely prevented the change in distribution of β chain radioactivity between supernatant and ribosome fractions caused by human β chains. It is suggested that, during rabbit hemoglobin synthesis, a pool of α chains, endogenous to the soluble phase of the reaction, aided in the release or solubilization of some of the completed rabbit β chains from the ribosome fraction. Human β chains, when added to the reaction, interacted with the α chains to prevent them from aiding in this release. Under these conditions some of the newly synthesized rabbit β chains were released as free chains, while others remained bound to the ribosomes. Addition of hemin to the reaction prevented the accumulation of ribosome-bound chains by aiding in their release. The enhanced release of newly synthesized β chains by soluble α chains or hemin may be part of a translational control mechanism in hemoglobin assembly; however, the presence of free 14C-β chains in the supernatants of reactions containing human β chains suggests that α chains were not required absolutely for this release.