Bohr effect in monomeric insect haemoglobins controlled by O2 off‐rate and modulated by haem‐rotational disorder

Abstract

The monomeric insect (Chironomus thummi thummi) haemoglobins CTT III and CTT IV show an alkaline Bohr effect. The amplitude of the Bohr effect curve of CTT IV is about twice as large as that of CTT III. In particular, at low pH a time‐dependent ‘slow’ decrease in p50 upon cyclic oxygenation/deoxygenation is observed which is larger if dithionite, instead of ascorbate, is the reducing agent. The decrease of p50 (increase in affinity) correlates with the ratio of haem‐rotational components exhibiting an increase of the ‘myoglobin‐like’ haemrotational component with high O2 affinity and high stability of the globin‐haem complex. The replacement of protohaem IX by mesohaem IX and deuterohaem IX, respectively, causes an increase in O2 affinity following the order: proto < meso proto‐III ≧ proto‐IX(y) CTT Hbs for low pH and proto‐IX(x) ≧ proto‐XIII > proto‐III ∼ proto‐IX(y) CTT Hbs for high pH] reflect the modulation of koff, O2 affinity, and Bohr effect exerted by specific contacts of the porphyrin substituents in position 1–4 with protein sites named A, B, C and D. The permutations of the porphyrin substituents in position 1–4 result in characteristic changes of the interactions at the protein sites A, B, C and D. The globin‐haem complex stability is largest and the koff value is smallest, if a vinyl, i.e. in proto‐IX(y) and proto‐III CTT Hbs, interacts with site C. The Bohr effect is largest if a vinyl, i.e. in proto‐IX(x) and proto‐III CTT Hbs, interacts with site B. Thus, the contact of vinyl with site B is an indicator of the t⇌r conformation transition. Copyright © 1986, Wiley Blackwell. All rights reserved