A growing number of proteins devoid of signal peptides have been demonstrated to be released through the non-classical pathways independent of endoplasmic reticulum and Golgi. Among them are two potent proangiogenic cytokines FGF1 and IL1α. Stress-induced transmembrane translocation of these proteins requires the assembly of copper-dependent multiprotein release complexes. It involves the interaction of exported proteins with the acidic phospholipids of the inner leaflet of the cell membrane and membrane destabilization. Not only stress, but also thrombin treatment and inhibition of Notch signaling stimulate the export of FGF1. Non-classical release of FGF1 and IL1α presents a promising target for treatment of cardiovascular, oncologic, and inflammatory disorders. Keywords non-classical secretion; FGF1; IL1α VARIETY OF NON-CLASSICALLY RELEASED PROTEINSThe familiar textbook scheme of protein secretion starts with the cotranslational protein translocation into the endoplasmic reticulum (ER). This translocation requires a molecular exit visa, a hydrophobic signal peptide located usually at the N-terminus of a secretable protein [Blobel, 1995]. After the protein is translocated through the ER membrane, its folding, transport, sorting, and covalent modifications occur in the ER and Golgi. Finally, the protein is released from the cell as a result of the fusion of an exocytotic vesicle with the cell membrane.

The conformational stability of the human acidic fibroblast growth factor (hFGF-1) is investigated using amide proton exchange and temperature-dependent chemical shifts, monitored by two-dimensional NMR spectroscopy. The change in free energy of unfolding (DeltaG(u)) of hFGF-1 is estimated to be 5.00 +/- 0.09 kcal.mol(-)(1). Amide proton-exchange rates of 74 residues (in hFGF-1) have been unambiguously measured, and the exchange process occurs predominately according to the conditions of the EX2 limit. The exchange rates of the fast-exchanging amide protons exposed to the solvent have been measured using the clean SEA-HSQC technique. The amide proton protection factor and temperature coefficient estimates show reasonably good correlation. Residues in beta-strands II and VI appear to constitute the stability core of the protein. Among the 12 beta-strands constituting the beta-barrel architecture of hFGF-1, beta-strand XI, located in the heparin binding domain, exhibits the lowest average protection factor value. Amide protons involved in the putative folding nucleation site in hFGF-1, identified by quench-flow NMR studies, do not represent the slow-exchanging core. Residues in portions of hFGF-1 experiencing high conformational flexibility mostly correspond to those involved in receptor recognition and binding.