A biophysical approach to the investigation of the properties of insulin and its receptor.
This thesis describes the properties of two hystricomorph
rodent insulins, those of the casiragua (Proechimys
guairae) and of the porcupine (Hystrix cristata). Studies of two bovine insulin analogues DAA and triphthaloyl insulins
are also presented.
Casiragua insulin has been extracted and purified and
its primary structure determined. The association properties
of casiragua insulin have been investigated by ultracentrifuge
studies and these show that it sediments as a monomer even
at high hormone concentration. Zinc binding studies reveal
that casiragua and porcupine insulins are unable to bind zinc
ions. Circular dichroism studies fully confirm the failure
of casiragua and porcupine insulins to associate and also
reveal changes in conformation compared with bovine insulin.
The biological properties of the insulins have been
examined by in vivo and in vitro bioassays and by receptor
binding assays on liver plasma membranes. These studies
reveal that casiragua and porcupine insulins have a low
potency in all mammalian tissues. Triphthaloyl insulin is
shown to be almost fully potent in hystricomorph tissues yet
of low activity in other mammals. On the basis of these
results it is suggested that hystricomorph insulin receptors
have undergone change. It is proposed that this, together
with changes in the structure of their insulins, has resulted
in a dimunition of the hypoglycaemic properties of these
insulins and a possible enhancement of other properties:
growth promoting properties for example.
The possible contribution of the B22-A21 ion pair to
the stability of insulin has been investigated by studieswith DAA insulin. Evidence is presented which suggests
that the ion pair is not as important in maintaining the
hormone in an active conformation as was previously thought.