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The question of whether a protein whose natural sequence is inverted adopts a stable fold is still under debate. We have determined the 2.1-Å crystal structure of the retro-GCN4 leucine zipper. In contrast to the two-stranded helical coiled-coil GCN4 leucine zipper, the retro-leucine zipper formed a very stable, parallel four-helix bundle, which now lends itself to further structural and functional studies. Since the early folding experiments by Anfinsen (1), it has been accepted that structure and function of a protein are determined by its amino acid sequence as read from the N terminus to the C terminus. But how does the structure change if the amino acid sequence of the protein is inverted? Inverted sequences are occasionally found in genomic DNA, but thus far, a native retro-protein has not been detected. Physicochemical properties that are related to the amino acid composition or the hydrophobicity profile should not be affected by the inversion of the sequence, supporting the idea that retro-sequences might fold into a native-like conformation. Modeling experiments suggested that reversal of the backbone direction may result in a topological mirror image of the native structure of the protein (2) or may produce the same topology as that of the parent protein (3, 4). Thus far, no structure elucidation of a retro-Lpeptide at atomic resolution has been reported.Coiled coils, including leucine zippers, consist of two to five intertwined ␣-helices and are frequently found in oligomeric proteins such as transcription factors as well as motility and structural proteins (5). We used the two-stranded coiled-coil domain of the yeast transcription activator GCN4 (6) to dimerize an artificial HIV enhancer-binding peptide, an operation that resulted in increased inhibition of HIV enhancer-controlled transcription (7). Because modeling studies suggested that the retro-sequence of the GCN4 leucine zipper also seemed to form a suitable dimerization module, a 35-residue retro-GCN4 was synthesized and characterized. Oxidized retro-leucine zipper extended with Cys-Gly-Gly, previously termed (r-LZ38) 2 (r-GCN4-p1Ј; Fig. 1A), crystallized and is now shown by x-ray structure analysis to fold into a parallel tetrameric coiled coil. Materials and MethodsUltracentrifugation. Sedimentation velocity experiments were performed with a Beckman-Spinco XL-A analytical ultracentrifuge. The peptide was dissolved in 20 mM Tris⅐HCl͞80 mM NaCl adjusted to pH 5.0, and measurements were made over a 10-to 100-M peptide concentration range. The centrifuge was operated at a speed of 50,000 rpm at 20°C. A partial specific volume, v 20 ϭ 0.74 cm 3 ⅐g Ϫ1, was calculated from the amino acid composition as was an estimate of the degree of hydration, d 1 ϭ 0.47 g water͞g protein. Sedimentation velocity traces were analyzed according to the method described in ref. 8, and a value of the sedimentation coefficient, s w,20 ϭ 1.77 Ϯ 0.5 Svedberg, was obtained by extrapolation to infinite dilution. The axial ratio was calculated as described in ref. 9.Crysta...

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Key words: carbohydrate-mediated antigenic epitope masking/ influenza virus glycoprotein/N-linked oligosaccharides

Based on secondary structure prediction rules and model building a neutral artificial 34-residue polypeptide with potential nucleic acid-binding activity was synthesised. This peptide and its covalent dimer showed strong interaction with cytidine phosphates and single-stranded DNA. The dimer had considerable ribonuclease activity with high preference for cleavage at the 3'-end of C.