title: Caveolin-1 is a competitive inhibitor of heme oxygenase-1 (HO-1) with heme: identification of a minimum sequence in caveolin-1 for binding to HO-1.
abstract: Heme oxygenase (HO) catalyzes the O(2)-dependent degradation of heme to biliverdin IXα, carbon monoxide (CO), and free ferrous iron through a multistep mechanism. Electrons required for HO catalysis in mammals are provided by NADPH-cytochrome P450 reductase. Recently, Kim et al. reported for the first time that HO, especially inducible HO-1, appears in caveolae and showed that caveolin-1, a principal isoform of the caveolin family, physically interacts with HO-1 [ Jung , N. H. et al. ( 2003 ) IUBMB Life 55 , 525 - 532 ; Kim , H. P. et al. ( 2004 ) FASEB J. 18 , 1080 - 1089 ]. In the present study, we confirmed by immunoprecipitation experiments that rat HO-1 and rat caveolin-1 (residues 1-101) directly interact with each other and that the HO-1 activity is inhibited by caveolin-1 (1-101). The 82-101 residues of caveolin-1 (CAV(82-101)), called the caveolin scaffolding domain, play essential roles in caveolin-related protein-protein interactions. The HO-1 activity is also inhibited by CAV(82-101) in a competitive manner with hemin, and a hemin titration experiment showed that CAV(82-101) interferes with hemin binding to HO-1. The enzyme kinetics and surface plasmon resonance experiments gave comparable K(i) and K(D) values of 5.2 and 1.0 μM for CAV(82-101), respectively, with respect to the interaction with HO-1. These observations indicated that CAV(82-101) and hemin share a common binding site within the HO-1 protein. The identified caveolin binding motif (FLLNIELF) of rat HO-1 is incomplete compared to the proposed consensus sequence. The affinity between HO-1 and CAV(82-101), however, was almost completely or remarkably eliminated by replacement of Phe(207) and/or Phe(214) with Ala, indicating that HO-1 binds to caveolin-1 via this motif. Among the peptide fragments derived from CAV(82-101), i.e., CAV(82-91), CAV(87-96), CAV(92-101), and CAV(97-101), CAV(92-101) and CAV(97-101) are able to inhibit the HO-1 activity to a similar extent; thus, the five-amino acid sequence (residues 97-101) is considered to be a minimum sequence for binding to HO-1.
authors: Department of Chemistry, Kurume University School of Medicine, Kurume 830-0011, Japan.
authors: Junichi
authors: J
authors: Taira
authors: Masakazu
authors: M
authors: Sugishima
authors: Yutaka
authors: Y
authors: Kida
authors: Eriko
authors: E
authors: Oda
authors: Masato
authors: M
authors: Noguchi
authors: Yuichiro
authors: Y
authors: Higashimoto
mesh_terms: D000818:Animals; D001665:Binding Sites; D051242:Caveolin 1 / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D006419:Heme Oxygenase (Decyclizing) / Q000037:antagonists & inhibitors* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011485:Protein Binding; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D020349:Surface Plasmon Resonance
chemical_list: D051242:Caveolin 1; D011994:Recombinant Proteins; D006419:Heme Oxygenase (Decyclizing); C491523:Hmox1 protein, rat