Mapping of heme-binding domains in soluble guanylyl cyclase beta1 subunit.
Soluble guanylyl cyclase (sGC) is activated upon the interaction of NO with heme in the sGC beta1 subunit. To identify the domains contributing to heme-binding, we constructed a series of deletion mutants of the beta1 subunit, and evaluated their heme-binding capability. Deletion mutants consisting of residues 1-120 [beta1(1-120)] and 80-385 [beta1(80-385)] were the shortest mutants exhibiting heme binding among the C-terminal and N-terminal-truncated mutants, respectively. The region common to both beta1(1-120) and beta1(80-385), i.e., residues 80-120, is therefore essential for heme binding, although the residues 341-385 play an auxiliary role in heme binding. Two deletion mutants, beta1(80-195) and beta1(60-195), which include only the essential region, exhibited strong heme binding and spectral properties similar to those of the nitrosyl complex of native sGC. Thus, these heme-binding core proteins may serve as model proteins for future studies on the tertiary structure of the nitrosyl complex of sGC.
Namiki
S
S
Department of Pharmacology, Graduate School of Medicine, University of Tokyo, Bunkyo-ku, Tokyo, 113-0033, Japan.
Hirose
K
K
Iino
M
M
D000818:Animals; D001665:Binding Sites; D001921:Brain / Q000201:enzymology / Q000378:metabolism; D004227:Dithionite / Q000378:metabolism; D006162:Guanylate Cyclase; D006418:Heme / Q000378:metabolism*; D007093:Imidazoles / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism; D011485:Protein Binding; D025941:Protein Interaction Mapping; D017434:Protein Structure, Tertiary; D021122:Protein Subunits; D051381:Rats; D018160:Receptors, Cytoplasmic and Nuclear / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion / Q000235:genetics; D000071756:Soluble Guanylyl Cyclase; D013057:Spectrum Analysis; D013879:Thioredoxins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D007093:Imidazoles; D021122:Protein Subunits; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D004227:Dithionite; D009569:Nitric Oxide; D006418:Heme; D013879:Thioredoxins; C029899:imidazole; D006162:Guanylate Cyclase; D000071756:Soluble Guanylyl Cyclase