Our lab is studying the structure of the rod and cone photoreceptor
phosphodiesterase that is central to the visual signaling pathway. The rod
enzyme contains a regulatory tandem GAF domain (GAFa-GAFb) and a catalytic
domain, and exists as a heterodimer (PDE6A and PDE6B).
There are currently no full length crystal structures for this catalytic
heterodimer, only a structure for the GAFa regulatory domain of cone PDE6C,
the GAF domain that binds cGMP noncatalytically.
The most homologous protein to PDE6 is the PDE that is abundant in vascular
smooth muscle and is the target for Viagra, PDE5 (a catalytic homodimer).
There is a dimeric structure for the PDE5 tandem GAF domains (GAFa-GAFb)
and for the isolated, monomeric catalytic domain, but no full-length
structures. cGMP binds to the GAFa domain of PDE5, the same as PDE6.
A more distantly related PDE, PDE2, has a crystal structure of the nearly
full-length, dimeric protein, but unfortunately the GAF domains of PDE2
occur in an opposite order compared to PDE5 and PDE6 (i.e., the cGMP
binding site is in GAFb).
See the attached diagram for a visual description of the domain
organization of the three PDEs.
We are trying to build a homology model for PDE6 heterodimer. The homology
model when PDE6 is threaded onto the PDE2 structure is poor. In contrast,
structural homology modeling of PDE6 GAF and catalytic domains using PDE5
as a template gives excellent results. Unfortunately, it is only in the
PDE2 structure that the molecular architecture of the nearly full-length
dimer has been determined, namely the region between the GAF and catalytic
domains and the relationship of the two catalytic domains.
Is there some way to rearrange (cut-and-past) the individual domains in
the sequence alignment to allow PDE5 to serve for the template for those
domains that have been determined, while still using the PDE2 template to
thread the PDE6 linker region and to define the relationship of the two
catalytic domains with each other?
Could you kindly give us some advice on how to do the alignment and build
the model? I would greatly appreciate your help.
Molecular, Cellular and Biomedical Sciences Department
University of New Hampshire
Durham, New Hampshire 03824