Auto-assemblaggio di nano-sistemi catalitici a base di peptidi e nanoparticelle d'oro.

The catalytic efficiency, mechanistic pathways, and structural complexity displayed by enzymes make them a tremendous source of inspiration for chemists involved in catalyst development[1, 2]. Nature has evolved enzymes as large multi-kilodalton complex structures in which even units that are remote from the actual active site may profoundly affect the activity of the enzyme[3]. The much lower complexity of artificial enzyme mimics may be an important reason for their typical modest performances with respect to enzymes. This awareness has led to an interest in catalysts based on multivalent scaffolds, such as dendrimers[4], micelles[5], and nanoparticles[6], with the idea of increasing the structural complexity of the synthetic system. A key challenge is straightforward access to synthetic catalysts that can match up to the size and complexity of enzymes. The necessity for multistep synthesis can be overcome by relying on self-assembly for the formation of the multivalent structure. In particular, the self-assembly of catalytic monolayers on the surface of gold nanoparticles (AuNPs) to give gold monolayer-protected clusters (AuMPCs) is emerging as an attractive strategy[7, 8]. Nonetheless, although they rely on self-assembly, the composition of self-assembled monolayers (SAMs) on Au NPs is typically still of rather low complexity[9]. Varying the surface composition by thiol clustering is a very difficult approach because this does not give a full control over the final composition, requires purification of each single NP system, and suffers from issues related to the characterization of mixed SAMs both in terms of composition and morphology. As a consequence, enormous efforts should be done to purify the different species coming from the synthesis. For that reason, in the recent years a new approach emerged by using nanoparticles not as direct tools, but as scaffolds to bind secondary molecules on. Rotello and coworkers were the first to realize this pioneering idea showing the attractiveness of cationic Au MPCs as a construction element for the development of innovative biosensors[10]. Inspired by those contributions, Prins et al. recently started to study the formation of heterofunctionalized multivalent structures relying on the self-assembly of small anionic peptides on the surface of Au MPCs[11, 12]. The results showed that it is possible to control the peptide surface composition simply exploiting the different affinities for the cationic monolayer. As a consequence, the low complexity of the surface can be overcome. This concept has been further developed during the PhD-project[13]. Based on the self-assembly of oligo-anions on a cationic surface, we went beyond the simple surface composition control and developed a true supramolecular nanoenzymatic system. It is composed by two fundamental elements: 1. Gold nanoparticles functionalized by alkyl thiols featuring a Trimethylammonium group in the Ω-position (8-Trimethylammonium octylthiol NR4+-AuNPs) thus generating a cationic surface; 2. Anionic oligopeptides that bind the nanoparticles’ surface. These feature a C-terminal tail with three Aspartic acid residues; an N-terminal tail composed by a variable number of Histidines (H0-H3); a central Tryptophane residue linking the two edges (Ac-HnWDDD-OH). The four negative charges coming from the C-terminal tail give an efficient binding, while Histidine and Tryptophane allow, respectively, catalysis and signal output for peptide concentration and binding measurements. Both of these constitutional elements are inefficient catalysts by themselves. Only when the peptides self-assemble on the cationic surface an active system is formed, which is able to accelerate the hydrolysis of N-CBz-(D)Phe-ONP by two orders of magnitude over the background. Importantly, the multivalent surface plays a crucial role in tuning the catalytic activity. The surface not only brings the substrate and catalyst in close proximity but also generates a microenvironment with an enhanced local pH that further activates the catalytic peptide. Given the supramolecular nature of the whole system and considering what has been written about the fine regulation of the surface composition, this system is highly adjustable simply by modulating the concentration of the constitutional elements that self-assemble on the surface. Once we obtained the lead system we started to investigate the intrinsic features that characterized it like the importance of the chemical structure of the substrate, the peptide catalyst and the cationic surface. The sequence of the peptide catalyst is very important for the efficiency of the catalytic system. Mutations on the H1 sequence (Ac-HWDDD-OH) altering its order and length showed that Triptophane has a subsidiary role in binding and its position should be next to the C-terminal tail. Histidine should occupy the N-terminal position because the presence of other residues in such position would reduce the catalytic efficiency of the Imidazole residue. If the sequence is extended by insertion of a Glycine residue between the Tryptophane and the C-terminal tail, we still observe a lower hydolysis rate, but not so drastic as the one observed by flipping the N-terminal sequence (steric hinderance). Like a natural enzyme, this system has specific requirements for the substrate that undergoes hydrolysis. SAR studies have been performed on substrate analogs with different Nα-protecting groups and side chains. The results showed that large and hydrophobic substrates have a higher affinity for the nanoparticles and are hydrolyzed faster. This is presumably due to interactions between those hydrophobic surfaces and the hydrophobic part of the monolayer. The aromatic substrates are favoured respect to the alkyl ones which emerges from the comparison between the kobs of N-CBz-Leu-ONP and N-CBz-(L)Trp-ONP: the latter is hydrolyzed much faster. The Nα-protecting group seems to have a crucial role in the substrate stabilization: diminishing its dimensions, thus diminishing hydrophobicity, results in a fast, spontaneous hydrolysis in simple buffer. A significant effort was made to improve the catalytic performances of the system through a supramolecular approach. In particular a hybrid system was prepared by self-assembling peptide H2 (Ac-HHWDDD-OH) and a library of non-catalytic peptides contemporarily on the monolayer surface (Ac-XXWDDD-OH, where XX are Leu, Phe, Ser and Arg in 42 possible combinations). The idea was that the second peptides would modulate the catalytic efficiency of the system. The results did not match the expectations which is presumably due to the fact that those peptides did not compensate the loss of the pH effect with additional interactions. Mutations on the H1 sequence (Ac-HWDDD-OH) by inserting identical flanking residues beside Histidine (Ac-XHXWDDD-OH) were studied with the scope of generating enantioselectivity. Although the variety of the flanking residues was large, (Leu, Phe, Ser, Tyr) no substrate enatioselectivity was observed. Despite this we had some experimental evidences that suggested that some enantioselectivity could be obtained by exploiting the binding of the substrate on the nanoparticles surface

Innovative Minichannel Condensers and Evaporators for Air Conditioning Equipment

The use of aluminum heat exchangers for refrigeration and air-conditioning equipment is very interesting since it allows to reduce weight and manufacturing costs while maintaining high performance. In this paper a two-phase heat transfer characterization of an innovative aluminum minichannel heat exchanger is presented. The heat exchanger (HX) is composed by rectangular channels with internal perforated turbolators. A special test section has been projected and realized in the Two Phase Heat Transfer Lab of the University of Padova in order to measure the heat transfer coefficient (HTC) on the refrigerant side during flow boiling and condensation. The test section has a single refrigerant channel with a perforated fin to make the minichannels. The test section is provided with 14 water flow modules installed at top and bottom of the refrigerant channel to promote boiling or condensation of the refrigerant. Therefore, the test section is made of seven different zones: each of them is equipped with 8 thermocouples to measure the wall temperature during the refrigerant phase change. The heat flow rate in each zone is calculated by an energy balance on the water side. Pressure transducers and thermocouples on the refrigerant side allow to determine the saturation temperature and thus the heat transfer coefficient of the refrigerant. The operating refrigerant used during tests is R410A. The particular scheme adopted for the test section enables to measure HTC at varying vapor quality and heat flow rate. Vaporization and condensation tests were carried out with different saturation temperatures, specific heat flow rate (from 40 to 150 kW/m2) and refrigerant mass flux (50÷150 kg/(m2 s) ). Data acquired have been compared with vaporization and condensation predictions from various correlations available in literature. This part of the work is very interesting since no data is available in the literature for such a geometry in vaporization nor in condensation. Therefore, the present paper will investigate the potential performance of these innovative minichannel heat exchangers as condensers and evaporators in air-conditioning equipment

label free fluorescence detection of kinase activity using a gold nanoparticle based indicator displacement assay

A straightforward fluorescence indicator-displacement assay (IDA) has been developed for the quantitative analysis of ATP→ADP conversion

Hyperoxia-induced changes in morphometric parameters of postnatal neurogenic sites in rat

In literature many works address the effects of hypoxia exposure on postnatal neurogenesis but few data are available about hyperoxia effects, although high oxygen concentrations are frequently used for ventilation of premature newborns. Thus, the aim of the present study was to compare with controls the morphometrical parameters of the main neurogenic sites (subventricular zone and dentate gyrus) in newborn Sprague-Dawley rats exposed to 60% or 95% oxygen for the first 14 postnatal days. Six rats were studied for each of the three groups. The unbiased quantitative method of the optical disector was applied to analyze neuronal densities, nuclear volumes, and total neuron numbers of the subventricular zone and hippocampal dentate gyrus. Apoptosis (terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling, TUNEL) and proliferation (Ki67) were also studied. The subventricular zone of newborn rats exposed to 95% hyperoxia showed statistically significant higher volume (mean value ± coefficient of variation: 0.40 ± 0.20 mm3) than subventricular zone of rats raised in normoxia (0.20 ± 0.11 mm3) or 60% hyperoxia (0.26 ± 0.18 mm3). Total neuron number was also significantly higher in 95% hyperoxia while neuronal densities did not reach statistically significant differences. TUNEL showed increased apoptotic indexes in hyperoxic rats. The percentage of proliferating KI67 positive cells was also higher in hyperoxia. The dentate gyrus granular layer of the normoxic rats showed higher volume (0.65 ± 0.11 mm3) than both the hyperoxic groups (60% hyperoxia: 0.39 ± 0.14 mm3; 95% hyperoxia: 0.36 ± 0.16 mm3). Total neuron numbers of hyperoxic dentate gyrus were also significantly reduced; neuronal densities were not modified. Hyperoxia-exposed rats also showed higher apoptotic and proliferating indexes in the dentate gyrus. Hyperoxia exposure in the first postnatal period may affect the main neurogenic areas (subventricular zone and dentate gyrus) increasing apoptosis but also inducing a certain reparative response consisting of increased proliferation. In particular, the increased volume of the subventricular zone may be ascribed to compensatory neurogenic response to the hyperoxic damage. Conversely, the decreased volume of the dentate gyrus granular layer could derive by a non sufficient neurogenic response to counterbalance the hyperoxic neuronal injury

L-citrulline is protective in hyperoxic lung damage and improves matrix remodelling and alveolarization

Moderate hyperoxia alters alveolar and vascular lung morphogenesis. Nitric oxide (NO) and matrix metalloproteinases (MMP) have a crucial role in the homeostasis of the matrix and bronchoalveolar structure and may be regulated abnormally by exposure to hyperoxia. Disruption of vascular endothelial growth factor (VEGF)-NO signaling impairs vascular growth and contributes to hyperoxia-induced vascular disease in bronchopulmonary dysplasia (BPD). We hypothesize that L-citrulline, by raising the serum levels of L-arginine and enhancing endogenous NO synthesis, might attenuate hyperoxia-induced lung injury in an experimental model of BPD. Neonatal rats (1 day old) were exposed to 60% oxygen or room air for 14 days and administered L-citrulline or a vehicle (sham). Lung morphometry were performed; Serum was tested for arginine level; Matrix metalloproteinases2 (MMP2) gene expression, VEGF gene and protein expression and endothelial NO synthase (eNOS) protein expression were compared. Mean linear intercept was higher in the hyperoxia and sham groups when compared with the room air (RA) and L-citr+hyperoxia treated group (p<0.02). Secondary crests number was higher in L-citrulline treated and RA when compared to hyperoxia and sham group (p<0.02). L-Arginine level rose in the L-citrulline-treated group (p<0.05). L-citrulline did not affect MMP2 gene expression, but it regulated the MMP2 active protein, which rose in bronchoalveolar lavage fluid (p<0.05), presumably due to a post-transductional effect. Compared with RA controls, hyperoxia significantly decreased VEGF and eNOS protein expression. At the same time, an increased lung VEGF gene and protein expression (p<0.05) were also seen in the rats treated with L-citrulline. We conclude that: (i) hyperoxia decreases growth and disrupts VEGF-NO signaling of lung; (ii) the main effects of L-citrulline are an increased serum level of arginine, as a promoter and a substrate of the nitric oxide synthase; and (ii) a better alveolar growth and matrix control than in hyperoxia-induced lung damage seems promising

Auto-assemblaggio di nano-sistemi catalitici a base di peptidi e nanoparticelle d'oro.

The catalytic efficiency, mechanistic pathways, and structural complexity displayed by enzymes make them a tremendous source of inspiration for chemists involved in catalyst development[1, 2]. Nature has evolved enzymes as large multi-kilodalton complex structures in which even units that are remote from the actual active site may profoundly affect the activity of the enzyme[3]. The much lower complexity of artificial enzyme mimics may be an important reason for their typical modest performances with respect to enzymes. This awareness has led to an interest in catalysts based on multivalent scaffolds, such as dendrimers[4], micelles[5], and nanoparticles[6], with the idea of increasing the structural complexity of the synthetic system. A key challenge is straightforward access to synthetic catalysts that can match up to the size and complexity of enzymes. The necessity for multistep synthesis can be overcome by relying on self-assembly for the formation of the multivalent structure. In particular, the self-assembly of catalytic monolayers on the surface of gold nanoparticles (AuNPs) to give gold monolayer-protected clusters (AuMPCs) is emerging as an attractive strategy[7, 8]. Nonetheless, although they rely on self-assembly, the composition of self-assembled monolayers (SAMs) on Au NPs is typically still of rather low complexity[9]. Varying the surface composition by thiol clustering is a very difficult approach because this does not give a full control over the final composition, requires purification of each single NP system, and suffers from issues related to the characterization of mixed SAMs both in terms of composition and morphology. As a consequence, enormous efforts should be done to purify the different species coming from the synthesis. For that reason, in the recent years a new approach emerged by using nanoparticles not as direct tools, but as scaffolds to bind secondary molecules on. Rotello and coworkers were the first to realize this pioneering idea showing the attractiveness of cationic Au MPCs as a construction element for the development of innovative biosensors[10]. Inspired by those contributions, Prins et al. recently started to study the formation of heterofunctionalized multivalent structures relying on the self-assembly of small anionic peptides on the surface of Au MPCs[11, 12]. The results showed that it is possible to control the peptide surface composition simply exploiting the different affinities for the cationic monolayer. As a consequence, the low complexity of the surface can be overcome. This concept has been further developed during the PhD-project[13]. Based on the self-assembly of oligo-anions on a cationic surface, we went beyond the simple surface composition control and developed a true supramolecular nanoenzymatic system. It is composed by two fundamental elements: 1. Gold nanoparticles functionalized by alkyl thiols featuring a Trimethylammonium group in the Ω-position (8-Trimethylammonium octylthiol NR4+-AuNPs) thus generating a cationic surface; 2. Anionic oligopeptides that bind the nanoparticles’ surface. These feature a C-terminal tail with three Aspartic acid residues; an N-terminal tail composed by a variable number of Histidines (H0-H3); a central Tryptophane residue linking the two edges (Ac-HnWDDD-OH). The four negative charges coming from the C-terminal tail give an efficient binding, while Histidine and Tryptophane allow, respectively, catalysis and signal output for peptide concentration and binding measurements. Both of these constitutional elements are inefficient catalysts by themselves. Only when the peptides self-assemble on the cationic surface an active system is formed, which is able to accelerate the hydrolysis of N-CBz-(D)Phe-ONP by two orders of magnitude over the background. Importantly, the multivalent surface plays a crucial role in tuning the catalytic activity. The surface not only brings the substrate and catalyst in close proximity but also generates a microenvironment with an enhanced local pH that further activates the catalytic peptide. Given the supramolecular nature of the whole system and considering what has been written about the fine regulation of the surface composition, this system is highly adjustable simply by modulating the concentration of the constitutional elements that self-assemble on the surface. Once we obtained the lead system we started to investigate the intrinsic features that characterized it like the importance of the chemical structure of the substrate, the peptide catalyst and the cationic surface. The sequence of the peptide catalyst is very important for the efficiency of the catalytic system. Mutations on the H1 sequence (Ac-HWDDD-OH) altering its order and length showed that Triptophane has a subsidiary role in binding and its position should be next to the C-terminal tail. Histidine should occupy the N-terminal position because the presence of other residues in such position would reduce the catalytic efficiency of the Imidazole residue. If the sequence is extended by insertion of a Glycine residue between the Tryptophane and the C-terminal tail, we still observe a lower hydolysis rate, but not so drastic as the one observed by flipping the N-terminal sequence (steric hinderance). Like a natural enzyme, this system has specific requirements for the substrate that undergoes hydrolysis. SAR studies have been performed on substrate analogs with different Nα-protecting groups and side chains. The results showed that large and hydrophobic substrates have a higher affinity for the nanoparticles and are hydrolyzed faster. This is presumably due to interactions between those hydrophobic surfaces and the hydrophobic part of the monolayer. The aromatic substrates are favoured respect to the alkyl ones which emerges from the comparison between the kobs of N-CBz-Leu-ONP and N-CBz-(L)Trp-ONP: the latter is hydrolyzed much faster. The Nα-protecting group seems to have a crucial role in the substrate stabilization: diminishing its dimensions, thus diminishing hydrophobicity, results in a fast, spontaneous hydrolysis in simple buffer. A significant effort was made to improve the catalytic performances of the system through a supramolecular approach. In particular a hybrid system was prepared by self-assembling peptide H2 (Ac-HHWDDD-OH) and a library of non-catalytic peptides contemporarily on the monolayer surface (Ac-XXWDDD-OH, where XX are Leu, Phe, Ser and Arg in 42 possible combinations). The idea was that the second peptides would modulate the catalytic efficiency of the system. The results did not match the expectations which is presumably due to the fact that those peptides did not compensate the loss of the pH effect with additional interactions. Mutations on the H1 sequence (Ac-HWDDD-OH) by inserting identical flanking residues beside Histidine (Ac-XHXWDDD-OH) were studied with the scope of generating enantioselectivity. Although the variety of the flanking residues was large, (Leu, Phe, Ser, Tyr) no substrate enatioselectivity was observed. Despite this we had some experimental evidences that suggested that some enantioselectivity could be obtained by exploiting the binding of the substrate on the nanoparticles surface.L’efficienza catalitica, i meccanismi e la complessità strutturale esibiti dagli enzimi sono da sempre stati un’inesauribile fonte d’ispirazione per i chimici impegnati nello sviluppo di nuovi catalizzatori[1, 2]. Gli enzimi si sono evoluti come grandi e complesse strutture di centinaia di migliaia di Dalton in cui subunità anche lontane dal sito attivo possono profondamente influire sull’attività dell’enzima medesimo[3]. La più bassa complessità strutturale dei sistemi catalitici artificiali basati sulla mimica enzimatica potrebbe essere un’importante ragione della loro efficienza tipicamente modesta. Questa consapevolezza ha diretto l’attenzione verso catalizzatori basati su scaffolds multivalenti come i dendrimeri[4], le micelle[5] e le nanoparticelle[6] con l’intento di aumentare la complessità dei suddetti sistemi sintetici. La sfida fondamentale è rappresentata dalla realizzazione semplice e diretta di catalizzatori sintetici che siano paragonabili in dimensioni e complessità agli enzimi. La necessità di sintesi laboriose può essere superata sfruttando l’auto-assemblaggio per l’ottenimento di strutture multivalenti. In particolare, l’auto-assemblaggio di monostrati catalitici sulla superficie di nanoparticelle d’oro (AuNPs) per dare clusters d’oro protetti da monostrato (Au MPCs) sta emergendo come un’efficacie strategia[7, 8]. Tuttavia, l’eccessiva omogeneità chimica della struttura derivante intrinsecamente dal processo di auto-assemblaggio dei tioli rimane un problema analogo all’eccessiva semplicità dei modelli non nano-derivati[9]. Tentativi di variegare la superficie delle nanoparticelle con tioli diversi mediante processi di clusterizzazione si sono dimostrati di difficile approccio; infatti, poiché la sintesi rimane basata sull’auto-assemblaggio, la composizione finale del sistema risulta troppo eterogenea in termini di decorazione delle superfici nanoparticellari. Ne conseguono enormi sforzi per la separazione e purificazione delle diverse specie. Per questa ragione negli ultimi anni si è fatto largo un nuovo approccio basato sulle nanoparticelle non come tools da impiegare in quanto tali, bensì come scaffolds dalla superficie omogenea ai quali far aderire molecole secondarie per ottenere sinergicamente l’effetto finale desiderato. Rotello e collaboratori sono stati tra i primi pionieri in questo ambito con lo sviluppo di innovativi biosensori basati su nanoparticelle d’oro funzionalizzate con tioli cationici[10]. Inspirandosi a questi lavori, recentemente, Prins et al. hanno sviluppato strutture etero-funzionalizzate, multivalenti basate sull’auto assemblaggio di piccoli ligandi sulla superficie di nanoparticelle d’oro[11, 12]. In questo modo è stato possibile dimostrare, con un approccio supramolecolare, come si possa finemente regolare la composizione della superficie delle nanoparticelle sfruttando le differenti affinità degli oligoanioni. Di conseguenza si è potuto superare in maniera semplice ed efficace il già citato problema dell’eccessiva omogeneità chimica legata ai sistemi supramolecolari e alle nanoparticelle d’oro funzionalizzate. Quest’idea è stata ulteriormente sviluppata durante l’internato di dottorato[13, 14]. Infatti, sfruttando la natura auto-assemblante degli oligo-anioni su sistemi nanoparticellari cationici, si è pensato di andare oltre il semplice controllo della composizione superficiale di ligandi e di realizzare un vero e proprio sistema nano-enzimatico artificiale di natura supramolecolare. Esso è composto da due elementi fondamentali: 1. Nanoparticelle d’oro passivate con tioli alchilici recanti in posizione Ω un gruppo trimetilammonico (8-Trimetilammonio ottiltiolo NR4+-AuNPs) in modo da generare una superficie cationica; 2. Oligo-peptidi anionici in grado di legarsi alla superficie delle nanoparticelle. Questi si caratterizzano per una coda C-terminale composta di tre residui di acido aspartico, per un’estremità N-terminale composta di un numero variabile d’Istidine (H0-H3), unite da un residuo di Triptofano (Ac-HnWDDD-OH). Le quattro cariche negative dei residui di Acido aspartico garantiscono un binding efficiente alla superficie cationica, mentre i residui d’Istidina e il Triptofano permettono, rispettivamente, la catalisi e la misurabilità del peptide stesso in termini di concentrazione e binding. Ciascuno dei due elementi preso singolarmente non è un efficiente catalizzatore. Infatti, solamente quando i peptidi si assemblano sulla superficie delle nanoparticelle, il sistema che ne scaturisce è in grado di accelerare l’idrolisi di un substrato modello come il p-Nitrofenil estere della N-CBz-(D)Fenilalanina di due ordini di grandezza rispetto al background. Oltretutto, la multivalenza della superficie nanoparticellare gioca un ruolo cruciale nella modulazione dell’attività catalitica non soltanto mantenendo substrato e catalizzatore in stretto contatto, ma generando un micro-ambiente dotato di un proprio pH locale che aumenta ulteriormente l’efficienza del sistema. Data la natura supramolecolare del sistema e ricordando quanto già visto per i lavori di Prins circa la regolazione della composizione della superficie cationica di nanoparticelle con oligoanioni, il sistema creato è finemente regolabile semplicemente dosando la quantità di componenti che si auto-assemblano sulla superficie delle nanoparticelle medesime. Una volta ottenuto il sistema lead si è voluto scendere nel dettaglio per comprendere le intime peculiarità che lo caratterizzavano come importanza della natura del substrato, importanza della natura dell’emi-catalizzatore peptidico ed importanza della natura della superficie cationica. La sequenza dell’emi-catalizzatore peptidico gioca un ruolo fondamentale nell’efficienza della catalisi con dei requisiti molto stringenti per quanto riguarda l’ordine dei singoli amminoacidi. Mutazioni a carico della sequenza H1 (Ac-HWDDD-OH) concernenti l’ordine e alla lunghezza hanno permesso di concludere che il Triptofano ha un ruolo accessorio nel binding e che la sua posizione deve essere adiacente alla coda anionica C-terminale. L’Istidina deve occupare l’estrema posizione N-terminale perché la presenza di amminoacidi in detta posizione ingombrerebbe il residuo imidazolico diminuendo l’efficienza catalitica del sistema. L’allungamento della sequenza, a parità di ordine, si traduce ancora una volta in una diminuzione dell’attività in ragione di un minore effetto del pH locale, ma con ripercussioni molto inferiori rispetto all’ingombro sterico. Analogamente ad un enzima naturale, tale sistema si è dimostrato avere dei precisi requisiti anche per quanto riguarda le caratteristiche dei substrati che ad esso si vanno a legare. Studi SAR sulla natura del substrato sono stati condotti utilizzando analoghi che presentassero modifiche alla natura e dimensioni del gruppo protettore dell’α-ammina e della catena laterale. I risultati ottenuti hanno permesso di concludere che, in generale, substrati più grandi e idrofobici hanno maggiore affinità per le nanoparticelle in ragione di un probabile nascondimento di superfici idrofobiche nel core alchilico. A parità d’idrofobicità le superfici aromatiche presentano una maggiore affinità e subiscono più facilmente idrolisi (si veda il caso di N-CBz-Leu-ONP e N-CBz-(L)Trp-ONP). Il gruppo protettore dell’α-ammina sembra rivestire un ruolo cruciale nella stabilizzazione del substrato: una riduzione delle sue dimensioni, con conseguente diminuzione dell’idrofobicità, si traduce in un’elevata tendenza all’idrolisi spontanea rendendo inutile il catalizzatore. Tentativi di migliorare la catalisi con approccio supramolecolare sono stati condotti auto-assemblando il sistema in maniera mista con il peptide H2 (Ac-HHWDDD-OH) e una libreria di peptidi (Ac-XXWDDD-OH, dove XX sono Leu, Phe, Ser e Arg nelle 42 combinazioni possibili) ritenuti essere modulatori dell’attività catalitica. I risultati ottenuti non sono stati in linea con le attese e questo probabilmente in relazione al fatto che il l’annullamento dell’effetto di pH non viene compensato dalle necessarie interazioni tra il sistema ed il substrato medesimo traducendosi in una diminuzione dell’attività. Mutazioni a carico della sequenza H1 mediante inserzione di coppie d’identici amminoacidi ai lati del residuo istidinico (Ac-XHXWDDD-OH) sono state condotte nel tentativo di ottenere un intorno chirale che permettesse una catalisi enantioselettiva. Nonostante la forte eterogeneità dei residui scelti (Leu, Phe, Ser, Tyr), il sistema non ha dato i risultati attesi. Dunque, a livello catalitico non sembra essere possibile discriminare gli enantiomeri, ma è auspicabile una discriminazione al momento del binding alla superfici cationica come ci hanno suggerito alcune evidenze sperimentali

Catalysis of Transesterification Reactions by a Self-Assembled Nanosystem

Histidine-containing peptides self-assemble on the surface of monolayer protected gold nanoparticles to form a catalytic system for transesterification reactions. Self-assembly is a prerequisite for catalysis, since the isolated peptides do not display catalytic activity by themselves. A series of catalytic peptides and substrates are studied in order to understand the structural parameters that are of relevance to the catalytic efficiency of the system. It is shown that the distance between the His-residue and the anionic tail does not affect the catalytic activity. On the other hand, the catalytic His-residue is sensitive to the chemical nature of the flanking amino acid residues. In particular, the presence of polar Ser-residues causes a significant increase in activity. Finally, kinetic studies of a series of substrates reveal that substrates with a hydrophobic component are very suitable for this catalytic system