Biology Reference
In-Depth Information
protein kinases, the cN-gated (CNG) channels and exchange-protein activated by
cAMP (EPAC). All PDEs contain a conserved catalytic site that interacts with cNs
and breaks them down into their respective 5
0
-nucleotides. Several PDEs (PDE 2, 5,
6, 10, and 11) contain cN-binding sites in their respective regulatory domains (Cote
2006
; Gross-Langenhoff et al.
2006
; Wu et al.
2004
; Zoraghi et al.
2005
; Handa
et al.
2008
; Martinez
2002a
; Martinez
2002b
; Martinez et al.
2008
); these sites
comprise ~120 amino acids known as GAFs [an acronym derived from the proteins
in which these domains were originally identified, i.e., cGMP-binding PDEs,
Ana-
baena
adenylyl cyclases, and
Escherichia coli
transcription factor FhlA (Aravind
and Ponting
1997
)]. For PDE 2, 5, and 6, cN binding to one of these GAFs regulates
catalytic site function, and it has been proposed that cN-binding GAFs in PDEs
could act to sequester cNs under appropriate conditions (Bender and Beavo
2006
;
Conti and Beavo
2007
; Corbin and Francis
1999
; Gopal et al.
2001
).
The respective cN-binding sites in GAFs are structurally and evolutionarily
distinct from the PDE catalytic sites and cN-binding sites in the cN-dependent
protein kinases, cN-regulated channels, the bacterial catabolite-gene activator pro-
tein (CAP), and EPACs (Bos
2006
; Charbonneau
1990
; Martinez et al.
2002a
,
b
;
Zoraghi et al.
2004
). The regulatory (R) and catalytic (C) subunits of cAMP-
dependent protein kinase (PKA), which combine to form the PKA holoenzymes,
are derived from four and three genes, respectively. Any R subunit (RI or RII),
which appear to exist as homodimers, can interact with any C subunit (C
a
,C
b
,
or C
g
) in mammalian tissues (Francis and Corbin
1999
). EPACI and II, which are
regulated by cAMP binding, are derived from two genes (Bos
2006
), and cGMP-
dependent protein kinases (PKGI and PKGII) are derived from two genes with
alternative splicing of the PKGI mRNA to produce two isoenzymes (PKGI
a
and
PKGI
b
) (Uhler
1993
; Wernet et al.
1989
). PKGs appear to always exist as homo-
dimers so it is predicted that there are only three PKGs in mammalian tissues. The
exceptionally diverse characteristics of the PDEs provide excellent potential for
development of selective inhibitors for these targets, but in most instances, their
similarities continue to confound development of such inhibitors.
The allosteric cN-binding sites on PDEs 2, 5, 6, and 11 preferentially bind
cGMP, although the sites on PDE2 can interact with cAMP with reasonable affinity
(Martinez et al.
2002a
,
b
; Wu et al.
2004
; Zoraghi et al.
2005
; Cote
2006
); the
allosteric cN-binding site in PDE10 tightly binds cAMP (Handa et al.
2008
). When
compared with other cN-binding sites, these sites are formed by a tight binding
pocket; in PDEs 5 and 6, this pocket rigorously selects for cGMP versus cAMP and
against substituents introduced in cGMP analogs (Huang et al.
2004
; Martinez
2002; Thomas et al.
1992
; Wu et al.
2004
). These characteristics make them
excellent targets for drugs that would impact the function of these PDEs. In PDEs
5 and 6, the allosteric site excludes PDE inhibitors. However, in PDE2, low
concentrations of 3-isobutyl-1-methylxanthine (IBMX) or papaverine stimulate
catalytic activity (Yamamoto et al.
1983
), although it is now unclear whether this
stimulation occurs through interaction with the allosteric cN-binding site or effects
mediated via partial occupation of the catalytic site (Pandit et al.
2009
). If this effect
is mediated through the allosteric cN-binding site, it lends some promise for
