Activation of soluble guanylate cyclase by NO-hemoproteins involves NO-heme exchange. Comparison of heme-containing and heme-deficient enzyme forms.
The mechanism of activation of soluble guanylate cyclase purified from bovine lung by high molecular weight, nitrosyl-hemoprotein complexes is reported. Heme-containing, heme-deficient, and heme-reconstituted forms of guanylate cyclase were studied. Nitric oxide (NO) and nitroso compounds activated heme-containing and heme-reconstituted enzymes (over 50-fold), with an accompanying shift in the Soret absorption peak from 431 to 398 nm, but failed to activate or alter the spectral characteristics of heme-deficient enzyme. In contrast, preformed NO-hemoprotein complexes as well as low molecular weight NO-heme activated all forms of guanylate cyclase. Heme-deficient guanylate cyclase was first reacted with excess amounts of NO-hemoglobin, NO-myoglobin, or NO-catalase and then rapidly separated from the NO-hemoprotein by column chromatography. Spectrophotometric analysis indicated that the NO-heme moiety was transferred from each of the NO-hemoproteins to heme-deficient guanylate cyclase. Approximately 1 mol of NO-heme was bound per mol of holoenzyme and the specific activity of this enzyme form was over 50-fold greater than that of unreacted, heme-deficient enzyme. NO-heme was tightly bound to guanylate cyclase as no transfer of enzyme-bound NO-heme to apohemoglobin was evident. Enzyme activated by NO-hemoproteins closely resembled, kinetically, that activated by NO or NO-heme. In contrast, reactions between heme-deficient guanylate cyclase and hemoproteins did not result in heme transfer, whereas heme alone rapidly reconstituted the enzyme. These observations indicate that soluble guanylate cyclase can be readily reconstituted with, and thereby activated by, NO-heme through an exchange reaction with NO-hemoproteins.
Ignarro
L J
LJ
Adams
J B
JB
Horwitz
P M
PM
Wood
K S
KS
D000818:Animals; D002374:Catalase / Q000494:pharmacology; D002417:Cattle; D004789:Enzyme Activation / Q000187:drug effects; D006162:Guanylate Cyclase / Q000378:metabolism*; D006418:Heme / Q000378:metabolism*; D006420:Hemeproteins / Q000494:pharmacology*; D006454:Hemoglobins / Q000494:pharmacology; D008168:Lung / Q000201:enzymology; D008970:Molecular Weight; D009211:Myoglobin / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism / Q000494:pharmacology*; D009603:Nitroso Compounds / Q000494:pharmacology*; D013053:Spectrophotometry
D006420:Hemeproteins; D006454:Hemoglobins; D009211:Myoglobin; D009603:Nitroso Compounds; D009569:Nitric Oxide; D006418:Heme; D002374:Catalase; D006162:Guanylate Cyclase