Soluble guanylyl cyclase is a heterodimer consisting of an alpha and b
eta subunit and stimulation occurs upon binding of NO to a prosthetic
group. Little is known about the localization of catalytic and regulat
ory domains within the subunits of soluble guanylyl cyclase. We used d
eletion mutagenesis to identify the regions of alpha(1) and beta(1) su
bunits that are responsible for cGMP production or NO-heme-mediated ac
tivation. The amino terminus of the beta(1) subunit was necessary for
NO stimulation since deletion of the 64 NH2-terminal amino acids resul
ted in a mutant with intact basal activity but complete loss of NO act
ivation. The amino terminus of the alpha(1) subunit also appeared to b
e essential for NO sensitivity since deletion of 131 NH2-terminal amin
o acids of alpha(1) led to markedly reduced NO activation. These resul
ts suggest that NH2-terminal regions of alpha(1) and beta(1) are invol
ved in NO-heme-mediated signal transduction. The NH2 terminally trunca
ted beta(1) subunit exerted a dominant negative effect exclusively on
the NO-stimulated activity of the wild type enzyme, further underlinin
g that the regulatory domain is located within the NH2 terminus of the
enzyme, Aside from the structural implications, the mutant represents
a powerful tool to investigate nitric oxide-sensitive signaling pathw
ays. Coexpression of the COOH-terminal halves of alpha(1) and beta(1)
were sufficient for basal cGMP production while either of the halves e
xpressed alone was inactive. Therefore the COOH-terminal regions appea
r to contain sufficient information for dimerization and basal enzymat
ic activity. Thus, we provide the first evidence that the regulatory a
nd catalytic properties of soluble guanylyl cyclase can be attributed
to different regions of the subunits and that the catalytic domain can
be functionally expressed separately from the NH2-terminal regulatory
domain. Taken together with findings on the membrane bound enzyme for
m, guanylyl cyclases, appear to resemble fusion proteins where differe
nt regulatory domains have been joined with a common cGMP-forming segm
ent.