The SH3 domain is a well characterized small protein module with a simple fold found in many proteins. At acid pH, the SH3 domain (PI3-SH3) of the p85α subunit of bovine phosphatidylinositol 3-kinase slowly forms a gel that consists of typical amyloid fibrils as assessed by electron microscopy, a Congo red binding assay, and x-ray fiber diffraction. The soluble form of PI3-SH3 at acid pH (the A state by a variety of techniques) from which fibrils are generated has been characterized. Circular dichroism in the far- and near-UV regions and ¹ H NMR indicate that the A state is substantially unfolded relative to the native protein at neutral pH. NMR diffusion measurements indicate, however, that the effective hydrodynamic radius of the A state is only 23% higher than that of the native protein and is 20% lower than that of the protein denatured in 3.5 M guanidinium chloride. In addition, the A state binds the hydrophobic dye 1-anilinonaphthalene-8-sulfonic acid, which suggests that SH3 in this state has a partially formed hydrophobic core. These results indicate that the A state is partially folded and support the hypothesis that partially folded states formed in solution are precursors of amyloid deposition. Moreover, that this domain aggregates into amyloid fibrils suggests that the potential for amyloid deposition may be a common property of proteins, and not only of a few proteins associated with disease.

Escherichia coli microcin C7 (MccC7) is an antibiotic that inhibits protein synthesis in vivo. It is a heptapeptide containing unknown modifications at the N and C termini (García-Bustos, J. F., Pezzi, N., and Méndez, E. (1985) Antimicrob. Agents Chemoth. 27, 791-797). The chemical structure of MccC7 has been characterized by use of ¹H homonuclear and heteronuclear (C, N, P) nuclear magnetic resonance spectroscopy as well as mass spectrometry (1177 ± 1 Da). The heptapeptide Met-Arg-Thr-Gly-Asn-Ala-Asp is substituted at the N terminus by a N-formyl group. The C-terminal substituent consists of the phosphodiester of 5′-adenylic acid and n-aminopropanol (AMPap), which is linked via the phosphorus atom to an amide group, thus forming a phosphoramide. The main chain carbonyl of the C-terminal aspartic acid residue is connected via this amide bond to the modified nucleotide unit. MccC7 and the peptide unit inhibit protein translation in vitro while a synthetic analog of the AMPap substituent is not active. Neither the peptide nor the AMPap molecule has an effect on the growth of MccC7-sensible cells. Our results strongly suggest that the peptide is responsible for MccC7 antibiotic activity while the C-terminal substituent is needed for MccC7 transport. Implications of the structure determined in this work for MccC7 synthesis and mode of action are discussed.

Resistance of Aspergillus fumigatus conidia to desiccation and their capacity to reach the alveoli are partly due to the presence of a hydrophobic layer composed of a protein from the hydrophobin family, called RodA, which covers the conidial surface. In A. fumigatus there are seven hydrophobins (RodA–RodG) belonging to class I and III. Most of them have never been studied. We constructed single and multiple hydrophobin-deletion mutants until the generation of a hydrophobin-free mutant. The phenotype, immunogenicity, and virulence of the mutants were studied. RODA is the most expressed hydrophobin in sporulating cultures, whereas RODB is upregulated in biofilm conditions and in vivo Only RodA, however, is responsible for rodlet formation, sporulation, conidial hydrophobicity, resistance to physical insult or anionic dyes, and immunological inertia of the conidia. None of the hydrophobin plays a role in biofilm formation or its hydrophobicity. RodA is the only needed hydrophobin in A. fumigatus, conditioning the structure, permeability, hydrophobicity, and immune-inertia of the cell wall surface in conidia. Moreover, the defect of rodlets on the conidial cell wall surface impacts on the drug sensitivity of the fungus.