Unique structure of Ascaris suum b5-type cytochrome: an additional alpha-helix and positively charged residues on the surface domain interact with redox partners.
Cytochrome b5 of the body wall of adult Ascaris suum, a porcine parasitic nematode, is a soluble protein that lacks a C-terminal membrane-anchoring domain, but possesses an N-terminal pre-sequence of 30 amino acids. During the maturation of cytochrome b5, the N-terminal pre-sequence is proteolytically cleaved to form the mature protein of 82 amino acid residues. A. suum cytochrome b5 is a basic protein containing more lysine residues and exhibiting a higher midpoint redox potential than its mammalian counterparts. We developed an expression system for the production of the recombinant nematode cytochrome b5, which is chemically and functionally identical with the native protein. Using this recombinant protein, we have determined the X-ray crystal structure of A. suum cytochrome b5 at 1.8 A (1 A=0.1 nm) resolution, and we have shown that this protein is involved in the reduction of nematode body-wall metmyoglobin. The crystal structure of A. suum cytochrome b5 consists of six alpha-helices and five beta-strands. It differs from its mammalian counterparts by having a head-to-tail disulphide bridge, as well as a four-residue insertion in the vicinity of the sixth ligating histidine, which forms an additional alpha-helix, alpha4A, between helices alpha4 and alpha5. A. suum cytochrome b5 exists predominantly as a haem-orientation B isomer. Furthermore, the haem plane is rotated approx. 80 degrees relative to the axis formed by haem-Fe and N atoms of the two histidine residues that are ligated to haem-Fe. The charge distribution around the haem crevice of A. suum cytochrome b5 is remarkably different from that of mammalian cytochrome b5 in that the nematode protein bears positively charged lysine residues surrounding the haem crevice. Using immunohistochemistry, we found that A. suum cytochrome b5 is present in the nematode hypodermis. Based on this histochemical and structural information, the physiological function of A. suum cytochrome b5 and its interaction with nematode metmyoglobin can be hypothesized.
Yokota
Takehiro
T
Science and Technology Office, Yokohama Center, Mitsubishi Chemical Corporation, 1000 Kamoshida-cho, Aoba, Yokohama 227-8502, Japan.
Nakajima
Yoshitaka
Y
Yamakura
Fumiyuki
F
Sugio
Shigetoshi
S
Hashimoto
Muneaki
M
Takamiya
Shinzaburo
S
D000595:Amino Acid Sequence; D000818:Animals; D017165:Ascaris suum / Q000737:chemistry* / Q000378:metabolism; D001665:Binding Sites; D015786:Cytochromes b5 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D015801:Helminth Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D017386:Sequence Homology, Amino Acid; D055672:Static Electricity
D015801:Helminth Proteins; D015786:Cytochromes b5