Increased oxygen affinity with normal heterotropic effects in hemoglobin Loire [α88(F9)Ala→Ser]

Abstract

Increased homotropic allosteric effect, while maintaining normal heterotropic effects, was observed in hemoglobin Loire. The oxygen binding curves, at equilibrium, and the kinetic measurements demonstrated that the substitution of a88(F9) Ala for a Ser results in increased oxygen affinity and decreased ns0 value. The function of the residues involved in the Bohr effect or in the regulation by 2,3-bisphosphoglycerate is not altered. The effects of bezafibrate, which binds specifically to the a chains, was similar to that observed in Hb A. The functional properties of Hb Loire may be explained by a slight displacement of some key residues of the Cterminal region of the a chain destabilizing the T structure.

NMR-based Binding Screen and Structural Analysis of the Complex Formed between α-Cobratoxin and an 18-Mer Cognate Peptide Derived from the α1 Subunit of the Nicotinic Acetylcholine Receptor fromTorpedo californica

“…Briefly, the peptide was expressed as a ketosteroid isomerase fusion protein using plasmid pET 31b(ϩ) in standard M9 medium and Escherichia coli strain BL21 (DE3) (Novagen) with the replacement of 15 NH 4 Cl for NH 4 Cl to uniformly enrich the peptide with 15 N. The peptide was cleaved from the fusion protein by reaction with cyanogen bromide, and it was then purified by reverse phase-high performance liquid chromatography. As before, the C-terminal homoserine lactone form of the ␣18-mer was used for the NMR studies described below.…”

“…X-ray crystal structures and NMR solution structures of several different ␣-neurotoxins reveal a conserved structural pattern consisting of a core of disulfides with three fingers or loops (3)(4)(5)(6)(7)(8)(9)(10)(11)(12). Short ␣-neurotoxins have four core disulfide bonds, whereas the long ␣-neurotoxins have, in addition, a fifth disulfide in the tip of loop II.…”

Purification and characterization of two platelet-aggregation inhibitors, named angustatin and H-toxin TA2, from the venom of Dendroaspis angusticeps

“…H-toxin TA 2 consisted of 60 amino acid residues and had a calculated molecular weight of The experimental conditions were as follows: a TSK ODS 80Ts column (4.6 Â 250 mm) was used with 0.1% trifluoroacetic acid as solvent A and 80% acetonitrile-20% solvent A as solvent B. Elution was performed with a linear gradient of 0.5% solvent B/min from 29% to 48% for 40 min. (Labhardt et al, 1988), and toxin TA 2 from D. angusticeps venom (Viljoen and Botes, 1974), as well as snake venom disintegrins such as echistatin from Echis carinatus venom (Dennis et al, 1990), trigramin from Trimeresurus (Protobothrops) gramineus venom (Dennis et al, 1990), barbourin from Sistrurus miliarius barbouri venom (Scarborough et al, 1991), kistrin from Agkistrodon rhodostoma venom (Dennis et al, 1990), and bitistatin from Bitis arietans venom (Shebuski et al, 1989). As shown in Fig.…”

Structure of fasciculin 2 from green mamba snake venom: evidence for unusual loop flexibility

“…These molecules, that have molecular weights close to 7000 Da, can bind to their target with high affinity, have no catalytic function and are functionally diversified. Numerous crystallographic analyses (Low et al, 1976;Tsernoglou & Petsko, 1976;Love & Stroud, 1986;Rees et al, 1987;Betzel et al, 1991;le Du, Marchot, Bougis & Fontecilla-Camps, 1992;Nickitenko, Michailov, Betzel & Wilson, 1993) as well as NMR studies (Labhardt, Hunziker-Kwik & Wtithrich, 1988;Steinmetz et al, 1988;Laplante et al, 1990;Yu, Lee, Chuang, Shei & Wang, 1990;Oswald et al, 1991 ;Brown & Wtithrich, 1992;Zinn-Justin et al, 1992;Goiovanov, Lomize, Arseniev, Utkin & Tsetlin, 1993;O'Connell, Bougis & Wi.ithrich, 1993) have shown that all these toxins are constituted of a dense core typically containing four disulfide bridges, and of three long loops (called I, II and III) emerging from the core, like the three central fingers of a hand. They all contain two ,8-sheets: a three-stranded sheet comprising loop II and one segment of loop II1, and a two-stranded one involving loop I.…”