α‐Helical assembly of biologically active peptides and designed helix bundle protein

Abstract

The formation of alpha-helical assembly by complexing biologically active peptides with de novo designed protein is described. The de novo designed protein described here is a cystine-linked 4-helix bundle protein constructed with 80 amino acid residues and forms a hydrophobic core region surrounded by 4 helices in an aqueous solution. The biologically active peptides, such as melittin and human growth hormone releasing factor, contain the sequences that are able to form amphiphilic helices. These peptides alone do not form the alpha-helix structure in a diluted solution with low ion strength. But on mixing with the designed helix bundle protein, the peptides are strongly bound to the protein with the induction of alpha-helical structure in the biologically active peptides. The content of induced alpha-helix is in accord with that estimated from the amphiphilic sequence. The results mean that a novel architecture composed of alpha-helices is formed. Fluorescent and temperature-scanning measurement revealed that the alpha-helical assembly is constructed with hydrophobic interaction. Also, it is shown by means of fluorescence depolarization that the assembly has a compact globular form corresponding to 1:1 complex.

Elucidating the Folding Problem of Helical Peptides using Empirical Parameters. II†. Helix Macrodipole Effects and Rational Modification of the Helical Content of Natural Peptides

“…1989; Greenfield et al, 1994;Jarvis et al, 1994;Jimenez et al, 1987Jimenez et al, , 1988Jimenez et al, , 1992Jimenez et al, , 1993Jimenez et al, , 1994Kemmink & Creighton, 1993;Kuroda, 1993;Mammi et al, 1992;Maulet & Cox, 1983;McDowell et al, 1985;Morii et al, 1994;Morton et al, 1994;Munier et al, 1993;Muñoz & Serrano, 1994, 1995Musco et al, unpublished results;Peñ a et al, 1989;Pintar et al, 1994;Precheur et al, 1991;Reed et al, 1992;Shin et al, 1993a,b;Siligard et al, 1991;Terzi et al, 1994;Waltho et al, 1993;Yumoto et al, 1993;Zhang et al, 1993;this work), while the other 288 peptides correspond to designed sequences Bradley et al, 1990;Chakrabartty et al, 1993bChakrabartty et al, , 1994DeGrado et al, 1985;Fairman et al, 1989Fairman et al, , 1991Forood et al, 1993;Ghadiri & Chong, 1990;Huyghues-Despointes et al, 1993a,b;Li & Deber, 1993;Lyu et al, 1989Lyu et al, , 1990Lyu et al, , 1991…”

“…To show that the success of our method is not due to an overfitting of the initial parameters to the peptide database used to refine them, it is necessary to carry out a blind test. For this purpose we set apart a number of peptides, which has been extended from that presented by Muñoz & Serrano (1994, 1995, corresponding mainly to protein fragments (57 peptides: Burke et al, 1991;Bruch et al, 1991;Doughty & Hu, 1993;Greenfield et al, 1991;Kuroda, 1993;Mammi et al, 1992;Maulet & Cox, 1983;McDowell et al, 1985;Morii et al, 1994;Munier et al, 1993;Muñoz & Serrano, 1994, 1995Pintar et al, 1994;Precheur et al, 1991;Reed et al, 1992;Siligard et al, 1991;Terzi et al, 1994;Yumoto et al, 1993;Zhang et al, 1993;Musco et al, unpublished results) but also designed peptides (56 peptides: DeGrado et al, 1985;Fairman et al, 1989;Ghadiri & Chong, 1990;Chakrabartty et al, 1994;Li & Deber, 1993;Scholtz et al, 1991b;Zhou et al, 1994b; the (AAQAA) n peptides described above). These peptides were obviously not analysed until the method was finally adjusted.…”

Melittin: a Membrane-active Peptide with Diverse Functions

“…These include apolipoproteins and peptide hormones (DeGrado et al 1982;Kaiser and Kezdy 1984;Morii et al 1994), signal peptides (Garnier et al 1980;Golding and O'Shea 1995), the envelope glycoprotein gp41 from the human immunodeficiency virus (Eisenberg and Wesson 1990;Rabenstein and Shin 1995), the poreforming peptide of pathogenic Entamoeba histolytica (Leippe et al 1991(Leippe et al , 1992 and the 25-residue presequence (p25) of subunit IV of yeast cytochrome oxidase (Clague and Cherry 1988). Interestingly, melittin exhibits sequence and structural similarities to a region of the tobacco mosaic virus coat protein (TMV CP), which is known to be critical for protein-protein and protein-RNA interactions (Marcos et al 1995).…”

Effect of micellar charge on the conformation and dynamics of melittin

“…However, despite the availability of a high-resolution crystal structure of tetrameric melittin in aqueous solution (Terwilliger and Eisenberg 1982), the structure of the membrane-bound form is not yet resolved by X-ray crystallography. Yet the importance of the membrane-bound form stems from the observation that the amphiphilic a-helical conformation of this hemolytic toxin in membranes resembles those of apolipoproteins and peptide hormones (Kaiser and Kezdy 1983;Morii et al 1994), signal peptides (Golding and O'Shea 1995), and the envelope glycoprotein gp41 from the human immunodeficiency virus (HIV) (Rabenstein and Shin 1995). Furthermore, understanding of melittin-membrane interactions assumes greater significance due to the recent finding that melittin mimics the N-terminal of HIV-1 virulence factor Nef1-25 (Barnham et al 1997).…”