Non-Canonical Ligand-Binding Events as Detected by NMR - Recent Developments in Biomolecular NMR - page 229

with

p 1f ~k 1f ½ C eq
p 1b ~k 1b
p 2f ~k 2f
p 2b ~k 2b

ð 10 : 21 Þ

If any of these probabilities is smaller than the inverse of the 'NMR
timescale', here 120 s 21 , there will be slow-exchange behavior. In the case
where excess IP6 is present, the concentration of the free cavity site becomes
exceedingly small. Hence p 1f , which is equal to p 1f for a two-site exchange
model (see Section 10.1) becomes very small as well, and this three-site system
is also in slow exchange.
This suggests that a model is needed in which the probability of free IP6
binding to Hb must remain finite, even when the cavity site is occupied by
another molecule of IP6.
A model for this situation is

k 1f

IzS / ?

IS

k 1b

k 2f

ISzC / ? ICzS ð 10 : 22 Þ

k 2b

k 3f

IzC / ?

IC

k 3b

with the requirement that

k 1f

k 1b x k 2f k 2b ~ k 3f

ð 10 : 23 Þ

k 3b

The extra site S on hemoglobin binds IP6 independently of the cavity site C.
Critical to this model is that IP6 can move from this site to the cavity site
without leaving the protein, e.g., by surface diffusion.
For IP6 this is the three-site exchange

p 1f p 2f

I / ? IS / ?

IC

p 1b p 2b

ð 10 : 24 Þ

p 3f

I / ? IC

p 3b