Disulphide-bond pattern and molecular modelling of the dimeric disintegrin EMF-10, a potent and selective integrin alpha5beta1 antagonist from Eristocophis macmahoni venom.
The disulphide-bond pattern of the heterodimeric disintegrin EMF-10, a potent and selective integrin alpha(5)beta(1) antagonist from Eristocophis macmahoni venom, was established by combination of amino-acid analysis, N-terminal sequencing and collision-induced dissociation by nanoelectrospray ionization quadrupole ion-trap MS of fragments isolated by reversed-phase HPLC after degradation of EMF-10 with oxalic acid. Each EMF-10 subunit contains four intrachain disulphide bonds. Two interchain cystine residues join the EMF-10 polypeptides. The intrachain linkages are conserved in monomeric disintegrins. A molecular model of EMF-10 was built using averaged NMR co-ordinates of flavoridin as a template. The active hairpin loops of the EMF-10 subunits occupy opposite locations at the ends of an elongated disulphide-bond ladder. In the EMF-10 model the N-terminal polypeptide of EMF-10B is close to the RGD-loop of the EMF-10A subunit, suggesting that the N-terminal region of the B-subunit could potentially influence the biological activity of the A-subunit.
Calvete
J J
JJ
Instituto de Biomedicina de Valencia, C.S.I.C., Jaime Roig 11, E-46010 Valencia, Spain. jcalvete@ibv.csic.es
Jürgens
M
M
Marcinkiewicz
C
C
Romero
A
A
Schrader
M
M
Niewiarowski
S
S
D000595:Amino Acid Sequence; D000818:Animals; D003545:Cysteine / Q000737:chemistry; D019281:Dimerization; D019483:Disintegrins / Q000737:chemistry* / Q000494:pharmacology*; D004220:Disulfides / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019815:Oxalic Acid / Q000737:chemistry; D011487:Protein Conformation; D017469:Receptors, Fibronectin / Q000037:antagonists & inhibitors*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014757:Viper Venoms / Q000737:chemistry* / Q000494:pharmacology*; D017819:Viperidae
D019483:Disintegrins; D004220:Disulfides; C122151:EMF10 protein, Eristocophis macmahoni; D017469:Receptors, Fibronectin; D014757:Viper Venoms; D019815:Oxalic Acid; D003545:Cysteine