Conversion of singly liganded carboxyhemoglobin to a rapidly reacting form

Experiments were earned out to study the reaction of the first molecule of CO with deoxyhemoglobin at 20°C in 0.1 M phosphate buffer at pH 7.0. A combined flow/laser photolysis method was used in which deoxyhemoglobin was mixed with enough CO to saturate between 2 and 9% of the heme sites and full photolysis was carried out at delays of between 0.3 and 100 s after mixing. The recombination reaction becomes progressively more rapid as this time delay is increased from 0.3 to 20 s indicating the formation of a rapidly reacting species from Hb(CO) 1 ). In simulations of this reaction both the on-rate constant and concentration of this rapidly reacting species were varied to obtain best fits to the data. For all samples 50% of the CO bound heme sites were converted within 20 s to a rapidly reacting form with a CO on-rate constant of 6 ± 1 μM -1 S -1 . It appears that this rapidly reacting species is not dimers so occurrence of a slow conformational change in the Hb(CO) 1 molecule is suggested as a possible explanation for these observations. The Two State model of Monod et al. appears to be unable to explain the formation of rapidly reacting material from Hb(CO) 1