Asymmetric inclusion by de novo designed proteins: Fluorescence probe studies on amphiphilic α‐helix bundles

Morii, Hisayuki; Ichimura, Kunihiro; Uedaira, Hatsuho

doi:10.1002/prot.340110206

Cited by 23 publications

(9 citation statements)

References 23 publications

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“…Previously the authors reported that the size of the helix bundle protein including the fluorescence probe is 3.0-3.5 nm (axial) X 3.0-4.0 nm (equatorial) . 4 The experimental results for the size of the peptide-protein assemblies is almost similar or slightly larger than this. Considering that the assemblies are 1 : 1 complexes and both the peptides and P4W have an a-helical structure, it is reasonable to assume that the assemblies have complexed helix bundle structures.…”

Section: Resultssupporting

confidence: 51%

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

et al. 1994

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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.

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Section: Resultssupporting

confidence: 51%

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

et al. 1994

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“…When this probe combines with various molten globular protein forms, smaller intensity increases, typically 10-to 150-fold, and smaller blue-shifts, typically to the 460-490 nm range, are observed. The blue-shifts and intensity increases are evidently the result of ANS becoming associated with the nonpolar, nonrigidly packed regions of the molten globular forms (Semisotnov et al, 1991;Morii et al, 1991;Handel et al, 1993).…”

Section: Discussionmentioning

confidence: 97%

“…The fluorescent molecule 1 -anilinonaphthalene-8-sulfonate (ANS) has been used to probe the molten globule state (Semisotnov et al, 1991;Morii et al, 1991;Handel et al, 1993). Classical studies show that, as the ANS probe moves from water into the heme binding pocket in apomyoglobin, there is an increase in intensity of about 245-fold and a blueshift from 515 nm in water to 454 nm when in the binding pocket (Stryer, 1965).…”

Section: Discussionmentioning

confidence: 99%

Structural changes accompanying chloroform-induced contraction of the filamentous phage fd

Roberts¹,

Dunker²

1993

Biochemistry

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Contact with a chloroform/water interface at 2 degrees C induces contraction of fd filamentous phage into rodlike I-forms; this contraction is accompanied by a decrease in the magnitude of circular dichroism spectral intensity near 222 nm and an increase near 210 nm. Comparisons with circular dichroism spectra of 100% helical poly-L-lysine and N-bromosuccinimide-oxidized fd phage indicate that the spectral change accompanying the fd to I-forms transition is due primarily to a change in the contributions from the single tryptophan (W26) of the major coat protein, with probably no significant change in the alpha-helix content. Further contraction of the rodlike I-forms to spherical S-forms at 25 degrees C is accompanied by a substantial general decrease in the magnitude of the ellipticity throughout the 230-210-nm region, which is indicative of a decrease in the alpha-helix content of the major coat protein. The similarity of the circular dichroism spectrum of S-forms with that of coat protein in detergents suggests that the S-form coat protein resembles the coat protein in lipid bilayers. The intrinsic fluorescence of W26 is quenched without red-shift (but perhaps a barely detectable blue-shift) following fd contraction to I-forms and S-forms. The accessibility of W26 to aqueous quenchers does not change significantly upon contraction. However, interaction with hydrophobic quenchers is dramatically altered in the contracted forms in a manner suggesting that the environment surrounding the tryptophan changes from native-protein-like in the fd filament to molten globule-like in the I-form rods and S-form spheroids. As discussed herein, certain features of these data support previous suggestions that chloroform-induced filamentous phage contraction may provide information about phage penetration and assembly in vivo.

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“…Thus, the ®nding of this new a-helical motif should provide a new insight into the folding pathway of a-helical proteins. Moreover, a-helical motifs have been extensively studied as prime candidates for attempts at de novo design of proteins with novel properties (DeGrado et al, 1989;Cohen & Parry, 1990;Hecht et al, 1990;Morii et al, 1991;Kamtekar et al, 1993). To this aim, the a-antiparallel helix scaffold present in the 3D structure of p8 MTCP1 should constitute a very useful tool: when compared to classical a-helical bundles, the disul-®de bonds should provide most of the stabilization energy, leaving a larger part of the protein structure available for mutations.…”

Section: Discussionmentioning

confidence: 99%

Solution structure of human p8MTCP1, a cysteine-rich protein encoded by the MTCP1 oncogene, reveals a new α-helical assembly motif

Barthe

Yang

Chiche

et al. 1997

Journal of Molecular Biology

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MTCP1 (for Mature-T-Cell Proliferation) is the first gene unequivocally identified in the group of uncommon leukemias with a mature phenotype. The three-dimensional solution structure of the human p8(MTCP1) protein encoded by the MTCP1 oncogene was determined by homonuclear proton two-dimensional NMR methods at 600 MHz. After sequence specific assignments, a total of 931 distance restraints and 57 dihedral restraints were collected. The location of the three previously unassigned disulfide bridges was determined from preliminary DIANA structures, using a statistical analysis of intercystinyl distances. The solution structure of p8(MTCP1) is presented as a set of 30 DIANA structures, further refined by restrained molecular dynamics using a simulated annealing protocol with the AMBER force field. The r.m.s.d. values with respect to the mean structure for the backbone and all heavy atoms for a family of 30 structures are 0.73(+/-0.28) and 1.17(+/-0.23) A, when the structured core of the protein (residues 5 to 63) is considered. The solution structure of p8(MTCP1) reveals an original scaffold consisting of three alpha helices, associated with a new cysteine motif. Two of the helices are covalently paired by two disulfide bridges, forming an alpha-hairpin which resembles an antiparallel coiled-coil. The third helix is oriented roughly parallel to the plane defined by the alpha-antiparallel motif and its axis forms an angle of approximately 60 degrees with respect to the main axis of this motif.

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Asymmetric inclusion by de novo designed proteins: Fluorescence probe studies on amphiphilic α‐helix bundles

Cited by 23 publications

References 23 publications

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

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

Structural changes accompanying chloroform-induced contraction of the filamentous phage fd

Solution structure of human p8MTCP1, a cysteine-rich protein encoded by the MTCP1 oncogene, reveals a new α-helical assembly motif

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