Abstract: A theoretical method is proposed to identify structural domains in proteins of known structures. It is based on the distribution of the local axes of the polypeptide chain. In particular, a statistical analysis is applied to the contributions of the local axes to the absolute writhing number, a topological property of a space curve resulting from the number of self- crossings in the curve projections onto a unit sphere. This finding supports the hypothesis that topological requirements should be satisfied in the process of protein folding and in the final organization of the tertiary structures.

Abstract: The histidine-containing protein HPr plays a central role in the phosphotransfer reaction that, in the bacterial phosphoenolpyruvate:sugar phosphotransferase system, leads to the phosphorylation of specific carbohydrates at the time of their translocation across the bacterial membrane. In Escherichia coli HPr is also able to phosphorylate the protein IIANtr, which is encoded by the rpoN operon and is involved in nitrogen regulation in bacteria. As the phosphoryl transfer reaction occurs concomitantly with the formation of a transient complex between HPr and IIANtr, the model of the P~HPr in complex with IIANtr (HPr~P~IIANtr) was built by in silico analysis. The model obtained is fully compatible with data describing the NMR chemical shifts relative to the interaction between HPr and IIAMtl, a protein which is structurally similar to IIANtr. The model shows that, due to good surface complementarity of the two proteins, intermolecular hydrogen bonds are formed by the invariant amino acids Arg17 of HPr and Arg57 of IIANtr. Other intermolecular interactions have hydrophobic character.

Abstract: The results of a 3 ns molecular dynamics simulation of the dodecamer duplex d(TATGGATCCATA)2 recognised by the Bam HI endonuclease are here presented. The DNA has been simulated as a flexible molecule using AMBER force field and the Ewald summation method which eliminates the undesired effects of truncation and permits to evaluate the full effects of electrostatic forces. The starting B conformation evolves toward a configuration quite close to that observed through X-ray diffraction in its complex with Bam HI. This configuration is fairly stable and the Watson-Crick hydrogen bonds are well maintained over the simulation trajectory. Hydration analysis indicates a preferential hydration for the phosphate than for the ester oxygens. Hydration shells in both the major and minor groove were observed. In both grooves the C-G pairs were found to be more hydrated than A-T pairs. The "spine of hydration" in the minor groove was clear. Water residence time are longer in the minor groove than in the major groove, although relatively short in both cases. No special long values are observed for sites where water molecules were observed by X-ray diffraction indicating that water molecules having an high probability to be located in a specific site are also fast exchanging.

Abstract: A neural network-based predictor is trained to distinguish the bonding states of cysteine in proteins starting from the residue chain. Training is performed using 2452 cysteine-containing segments extracted from 641 non homologous proteins of well resolved 3D structure. After a cross-validation procedure efficiency of the prediction scores as high as 72% when the predictor is trained using protein single sequences. The addition of evolutionary information in the form of multiple sequence alignment and a jury of neural networks increase the prediction efficiency up to 81%. Assessment of the goodness of the prediction with a reliability index indicates that more than 60% of the predictions have an accuracy level greater than 90%. A comparison with a statistical method previously described and tested on the same database shows that the neural network-based predictor is performing with the highest efficiency.

Abstract: Taste receptors have been studied less than those of other stimuli. However, the availability of many agonists and the practical relevance of sweeteners have stimulated indirect studies of the interaction of sweet agonists with their receptor and the development of general models of the sweet receptor active site. Most sweeteners are small molecular weight compounds but there are also sweet macromolecules, both synthetic and natural, i.e., sweet proteins. Do they interact with the same receptor of low molecular weight compounds? There are several sweet proteins: miraculin , monellin, thaumatin, curculin, mabinlin, pentadin and brazzein, but only three of them, i.e. thaumatin, monellin and brazzein, have been studied from a structural point of view. Multiple alignment of the sequences of sweet proteins shows no similarity. There is also no obvious similarity among the structures of thaumatin, monellin and brazzein. How can we identify the protein glucophores? We made the assumption that they are similar to those of low molecular weight compounds and that all sweet compounds interact with the same receptor. In fact, our model for the sweet receptor (Temussi et al., 1984, 1991) is consistent also with macromolecules since the active site is depicted as an open cavity with a flat bottom. When trying to explain the sweet taste of a protein it's natural to assume the existence of some kind of "sweet finger", i.e., a protruding structural element hosting one or more glucophores. We sought to identify sweet fingers in the three sweet proteins whose structure is known. Detailed structure comparison of all loops in the structures of thaumatin, monellin and brazzein by means of DALI shows that each protein hosts a likely sweet finger in which the spatial arrangement of three key residues (an aromatic a hydrogen bond donor and a hydrogen bond acceptor) is consistent with our model of the receptor active site.

Abstract: Abbiamo sviluppato una procedura (de Rinaldis et al., 1998) per calcolatore che consente di analizzare e confrontare superficie proteiche che siano associate a specifiche funzioni (es.: particolari abilita' di legame o di catalisi). Sulla base di un allineamento multiplo di strutture proteiche omofunzionali ed in analogia con il metodo dei profili per la ricerca di omologie di sequenza (Gribskov et al., 1987), il programma permette di realizzare un profilo 3D che puo' essere usato per fare ricerche nel PDB e selezionare proteine con particolari caratteristiche di superficie e funzionali. L'analisi del profilo 3D consente l'individuazione di residui conservati sulla superficie delle proteine sovrapposte e la definizione di determinanti strutturali associati a particolari funzioni biologiche. Questo metodo puo' inoltre essere utilizzato: i) per identificare strutture proteiche che, pur avendo fold differenti, abbiano una o piu' regioni di superficie con proprieta' chimiche e funzionali simili (evoluzione convergente); ii) come sistema esperto per la mutagenesi sito-specifica o iii) per il protein design. Stiamo utilizzando i profili 3D per analizzare i determinanti di superficie legati a sei motivi del database PROSITE: aminoacyl-transfer RNA sintetasi di classe II, perossidasi , dominio EF-hand di legame al calcio, sito di legame all'eme della famiglia citocromo c, sito di legame a nucleotidi del p loop, sito attivo delle aspartil proteasi eucariotiche e virali. Tali motivi di sequenza non sono in grado di riconoscere tutte e sole le sequenze proteiche associate alla loro funzione, ma selezionano anche falsi positivi e talvolta non selezionano tutte le sequenze cui e' associata la funzione loro assegnata. Nostro scopo e' indagare se invece esista e possa essere identificato un motivo di superficie specifico per ognuna delle funzioni analizzate. Nel caso del p loop, l'analisi dei risultati ha permesso considerazioni di particolare interesse biologico. L'applicazione di questo metodo all'intero database di strutture note potrebbe consentire la realizzazione di una banca dati di motivi di superficie capaci di identificare tutte e sole le strutture associate ad una specifica funzione, in quei casi in cui l'analisi della sola sequenza non lo consentirebbe. Lo sviluppo dei progetti di "structural genomics" rende molto piu' vasto l'insieme delle proteine cui applicare il metodo.