Canonical, deprotonated, or zwitterionic? II. A computational study on amino acid interaction with the TiO2(110) rutile surface: comparison with the anatase (101) surface.

The adsorption of 11 amino acids (Gly, Leu, Met, Phe, Ser, Cys, Glu, Gln, Arg, Lys, and His) on the TiO2(110) rutile surface is investigated adopting a theoretical approach, using the PBE-D2* functional as implemented in the periodic VASP code. The adsorption of the amino acids is considered in their canonical, deprotonated and zwitterionic forms. For all cases, the most stable adsorption mode adopts a bidentate (O,O) binding with surface undercoordinated Ti atoms, in agreement with previous experimental and computational studies using glycine as a test case. Such a binding mode is possible due to the surface morphology, because the Ti-Ti distances match very well with the carboxylic O-O distance. The most stable adsorption states are the deprotonated and the zwitterionic ones, the canonical one lying significantly above in energy. The relative stability between the deprotonated and the zwitterionic states results in a delicate trade-off among dative interactions (O, N, and S atoms of the amino acids with Ti atoms of the surface), H-bond interactions, dispersive forces and, to a lesser extent, steric hindrance of the amino acidic lateral chains. Finally, the difference in the amino acid adsorption between the (110) rutile and the (101) anatase surfaces is discussed both from the energetic and surface morphological standpoints, highlighting the larger reactivity of the rutile polymorph in adsorbing and deprotonating the amino acids compared with the anatase one.

Mannose-Binding Lectin-Deficient Donors Increase the Risk of Bacterial Infection and Bacterial Infection-Related Mortality After Liver Transplantation.

Mannose-binding lectin (MBL) is synthesized by the liver and binds to microbes. MBL2 gene polymorphisms produce intermediate/low/null or normal MBL serum levels (MBL-deficient or MBL-sufficient phenotypes, respectively). We aimed to evaluate the incidence and severity of infection, rejection, and survival within 1 year after liver transplantation (LT) according to donor and recipient MBL2 gene polymorphisms. A repeated-event analysis for infection episodes (negative binomial regression, Andersen-Gill model) was performed in 240 LTs. Four hundred twenty-eight infectious episodes (310 bacterial, 15 fungal, 65 cytomegalovirus [CMV]-related, and 38 viral non-CMV-related episodes) and 48 rejection episodes were recorded. The main bacterial infections were urinary (n = 82, 26%) and pneumonia (n = 69, 22%). LT recipients of MBL-deficient livers had a higher risk of bacterial infection (incidence rate ratio [IRR] 1.48 [95% confidence interval 1.04-2.09], p = 0.028), pneumonia (IRR 2.4 [95% confidence interval 1.33-4.33], p = 0.013), and septic shock (IRR 5.62 [95% confidence interval 1.92-16.4], p = 0.002) compared with recipients of MBL-deficient livers. The 1-year bacterial infection-related mortality was higher in recipients of MBL-deficient versus MBL-sufficient livers (65.8% vs. 56.1%, respectively; p = 0.0097). The incidence of rejection, viral, or fungal infection was similar in both groups. Recipient MBL2 genotype did not significantly increase the risk of bacterial infection. LT recipients of MBL-deficient livers have a higher risk of bacterial infection, pneumonia, septic shock, and 1-year bacterial infection-related mortality after LT.

Reactivity of Metal Carbenes with Olefins: Theoretical Insights on the Carbene Electronic Structure and Cyclopropanation Reaction Mechanism.

Present work addresses the reactivity of several phenyl-substituted metal-carbene complexes with 4-methylstyrene by means of density functional theory OPBE simulations. Different paths that lead to cyclopropanation were explored and compared to the olefin metathesis mechanism. For this purpose, we chose four different catalysts: (i) the Grubbs second-generation olefin metathesis catalyst, (ii) a Grubs second-generation-like complex, in which ruthenium is replaced by iron, and (iii) two iron carbene complexes (a piano stool and a porphyrin iron carbene) that experimentally catalyze alkene cyclopropanation. Results suggest that the nature of the applying mechanism is very sensitive to the coordination around the metal center and the spin state of the metal-carbene complex. Cyclopropanation by open-shell metal-carbene complexes seems to preferentially proceed through a two-step radical mechanism, in which the two C-C bonds are sequentially formed (path C). Singlet-state carbenes proceed either through a direct attack of the olefin to the carbene (path D) when the formation of the metallacycle is not feasible or through a reductive elimination from the metallacyclobutane when this intermediate is accessible both kinetically and thermodynamically (path B).

Hepatic Infiltrates in Operational Tolerant Patients After Liver Transplantation Show Enrichment of Regulatory T Cells Before Proinflammatory Genes Are Downregulated.

Immunosuppression can be discontinued from selected and stable patients after liver transplantation resulting in spontaneous operational tolerance (SOT), although the underlying mechanisms remain elusive. Thus, we analyzed serial liver biopsy specimens from adult liver recipients enrolled in a prospective multicenter immunosuppression withdrawal trial that used immunophenotyping and transcriptional profiling. Liver specimens were collected before the initiation of weaning, at the time of rejection, or at 1 and 3 years after complete drug discontinuation. Unexpectedly, the tolerated grafts developed portal tract expansion with increased T cell infiltration after immunosuppression withdrawal. This was associated with transient and preferential accumulation of CD4(+) FOXP3(+) cells and a trend toward upregulation of immune activation and regulatory genes, without signs of rejection. At the same time, no markers of endothelial damage or activation were noted. Portal infiltrates persisted at 3 years but were characterized by decreased expression of genes associated with chronic immunological damage. Further, SOT was not associated with a progressive liver fibrosis up to 5 years. These data suggest that SOT involves several mechanisms: a long-lasting local immune cell persistence with a transient regulatory T cells accumulation followed by a downregulation of immune-activated genes over years. These results have important implications for designs and follow-up of weaning trials.

Molecular Characterization of Acute Cellular Rejection Occurring During Intentional Immunosuppression Withdrawal in Liver Transplantation.

Acute cellular rejection occurs frequently during the first few weeks following liver transplantation. During this period, its molecular phenotype is confounded by peri- and postoperative proinflammatory events. To unambiguously define the molecular profile associated with rejection, we collected sequential biological specimens from 55 patients at least 3 years after liver transplantation who developed rejection during trials of intentional immunosuppression withdrawal. We analyzed liver tissue and blood samples obtained before initiation of drug withdrawal and at rejection, alongside blood samples collected during the weaning process. Gene expression profiling was conducted using whole-genome microarrays and real-time polymerase chain reaction. Rejection resulted in distinct blood and liver tissue transcriptional changes in patients who were either positive or negative for hepatitis C virus (HCV). Gene expression changes were mostly independent from pharmacological immunosuppression, and their magnitude correlated with severity of histological damage. Differential expression of a subset of genes overlapped across all conditions. These were used to define a blood predictive model that accurately identified rejection in HCV-negative, but not HCV-positive, patients. Changes were detectable 1-2 mo before rejection was diagnosed. Our results provide insight into the molecular processes underlying acute cellular rejection in liver transplantation and help clarify the potential utility and limitations of transcriptional biomarkers in this setting.

Liver Retransplantation in Patients With HIV-1 Infection: An International Multicenter Cohort Study.

Liver retransplantation is performed in HIV-infected patients, although its outcome is not well known. In an international cohort study (eight countries), 37 (6%; 32 coinfected with hepatitis C virus [HCV] and five with hepatitis B virus [HBV]) of 600 HIV-infected patients who had undergone liver transplant were retransplanted. The main indications for retransplantation were vascular complications (35%), primary graft nonfunction (22%), rejection (19%), and HCV recurrence (13%). Overall, 19 patients (51%) died after retransplantation. Survival at 1, 3, and 5 years was 56%, 51%, and 51%, respectively. Among patients with HCV coinfection, HCV RNA replication status at retransplantation was the only significant prognostic factor. Patients with undetectable versus detectable HCV RNA had a survival probability of 80% versus 39% at 1 year and 80% versus 30% at 3 and 5 years (p = 0.025). Recurrence of hepatitis C was the main cause of death in the latter. Patients with HBV coinfection had survival of 80% at 1, 3, and 5 years after retransplantation. HIV infection was adequately controlled with antiretroviral therapy. In conclusion, liver retransplantation is an acceptable option for HIV-infected patients with HBV or HCV coinfection but undetectable HCV RNA. Retransplantation in patients with HCV replication should be reassessed prospectively in the era of new direct antiviral agents.

Somatostatin therapy protects porcine livers in small-for-size liver transplantation.

Small-for-size (SFS) injury occurs in partial liver transplantation due to several factors, including excessive portal inflow and insufficient intragraft responses. We aim to determine the role somatostatin plays in reducing portal hyperperfusion and preventing the cascade of deleterious events produced in small grafts. A porcine model of 20% liver transplantation is performed. Perioperatively treated recipients receive somatostatin and untreated controls standard intravenous fluids. Recipients are followed for up to 5 days. In vitro studies are also performed to determine direct protective effects of somatostatin on hepatic stellate cells (HSC) and sinusoidal endothelial cells (SEC). At reperfusion, portal vein flow (PVF) per gram of tissue increased fourfold in untreated animals versus approximately threefold among treated recipients (p = 0.033). Postoperatively, markers of hepatocellular, SEC and HSC injury were improved among treated animals. Hepatic regeneration occurred in a slower but more orderly fashion among treated grafts; functional recovery was also significantly better. In vitro studies revealed that somatostatin directly reduces HSC activation, though no direct effect on SEC was found. In SFS transplantation, somatostatin reduces PVF and protects SEC in the critical postreperfusion period. Somatostatin also exerts a direct cytoprotective effect on HSC, independent of changes in PVF.

Should IFN-γ, IL-17 and IL-2 be considered predictive biomarkers of acute rejection in liver and kidney transplant? Results of a multicentric study.

Acute rejection (AR) remains a major challenge in organ transplantation, and there is a need for predictive biomarkers. In the present multicenter study, we prospectively examined a series of biomarkers in liver and kidney recipients. Intracellular expression of IFN-γ, IL-17 and IL-2 and IL-17 soluble production were evaluated both pre-transplantation and post-transplantation (1st and 2nd week, 1st, 2nd and 3rd month). 142 transplant patients (63 liver/79 kidney) were included in the study. Twenty-eight recipients (14 liver/14 kidney) developed AR. Pre- and post-transplantation intracellular expression of %IFN-γ(+) in CD4(+)CD69(+) and in CD8(+)CD69(+) and soluble IL17 identified liver and kidney transplant patients at high risk of AR. Pre-transplantation, %IL-2(+) in CD8(+)CD69(+) also identified kidney patients at high risk. We constructed pre- and post-transplantation risk prediction models, based on a composite panel of biomarkers, which could provide the basis for future studies and will be a useful tool for the selection and adjustment of immunosuppressive treatments.

Formaldehyde chemistry in cometary ices: the case of HOCH2OH formation.

Laboratory experiments devoted to simulate the chemistry occurring in interstellar and cometary ice analogues are of paramount importance to understand the formation of complex organic molecules that are detected throughout the universe. These laboratory simulations provide relevant hints on the fundamental physical and chemical steps associated with the increase of the molecular complexity in space and, moreover, give benchmark results for dedicated space missions. In the present work, we study the thermally promoted reactivity of H2O-dominated and D2O-dominated cometary ice analogues that contain various amounts of H2CO and NH3 by means of Fourier-transform infrared spectroscopy (FTIR), mass spectrometry and DFT calculations. Experimental measurements show that methyleneglycol (HOCH2OH) and D2-methyleneglycol (DOCH2OD, the corresponding isotopologue) are formed from the H2O- and D2O-dominated ices, respectively, only if ammonia is present. We also reported for the first time the mass spectrum of methyleneglycol and D2-methyleneglycol. B3LYP calculations have also been used to characterize the potential energy surface of the mechanistic steps associated with the formation of HOCH2OH as well as to simulate the IR spectrum of this compound. The fruitful interplay between theory and experiment has allowed us to elucidate the exact role of ammonia during the warming, which essentially stands for the formation and stabilization of the NH4(+)/OH(-) ion pair, thus enabling the OH(-) species to react with formaldehyde. The present results reproduce the heating of circumstellar ices in star formation regions and can be applied to the late thermal evolution of comets. In addition, the mass spectrum of methyleneglycol represents a benchmark for the analysis of the data coming from the ROSINA on-board instrument of the Rosetta mission.

Intracellular IFN-γ and IL-2 expression monitoring as surrogate markers of the risk of acute rejection and personal drug response in de novo liver transplant recipients.

Biomarker monitoring is needed in transplantation to reflect individual response to immunosuppressive drugs and graft outcome. We evaluated intracellular expression and soluble production of interferon-(IFN)-γ and interleukin-(IL)-2 as predictive biomarkers of acute rejection (AR) and personal drug response. Pharmacokinetic-pharmacodynamic profiles were determined in 47 de novo liver recipients treated with tacrolimus, mycophenolate mofetil and prednisone. Of the 47 patients, AR occurred in nine. There were no differences in drug concentrations between rejectors and non-rejectors. A pre-transplantation cut-off value of 55.80% for %CD8(+)-IFN-γ(+) identified patients at high risk of AR with a sensitivity of 75% and a specificity of 82%. In the first week post-transplantation, patients with a % inhibition for soluble IFN-γ, %CD8(+)-IFN-γ(+) and %CD8(+)-IL2(+) lower than 40% developed AR, showing low susceptibility to immunosuppressive drugs. Therefore, effector-T-cell response monitoring may help physicians to identify personal response to treatment and patients at high risk of AR.

Tauroursodeoxycholic acid affects PPARγ and TLR4 in Steatotic liver transplantation.

Numerous steatotic livers are discarded for transplantation because of their poor tolerance to ischemia-reperfusion (I/R). We examined whether tauroursodeoxycholic acid (TUDCA), a known inhibitor of endoplasmic reticulum (ER) stress, protects steatotic and nonsteatotic liver grafts preserved during 6 h in University of Wisconsin (UW) solution and transplanted. The protective mechanisms of TUDCA were also examined. Neither unfolded protein response (UPR) induction nor ER stress was evidenced in steatotic and nonsteatotic liver grafts after 6 h in UW preservation solution. TUDCA only protected steatotic livers grafts and did so through a mechanism independent of ER stress. It reduced proliferator-activated receptor-γ (PPARγ) and damage. When PPARγ was activated, TUDCA did not reduce damage. TUDCA, which inhibited PPARγ, and the PPARγ antagonist treatment up-regulated toll-like receptor 4 (TLR4), specifically the TIR domain-containing adaptor inducing IFNß (TRIF) pathway. TLR4 agonist treatment reduced damage in steatotic liver grafts. When TLR4 action was inhibited, PPARγ antagonists did not protect steatotic liver grafts. In conclusion, TUDCA reduced PPARγ and this in turn up-regulated the TLR4 pathway, thus protecting steatotic liver grafts. TLR4 activating-based strategies could reduce the inherent risk of steatotic liver failure after transplantation.

Applicability and results of Maastricht type 2 donation after cardiac death liver transplantation.

Maastricht type 2 donation after cardiac death (DCD) donors suffer sudden and unexpected cardiac arrest, typically outside the hospital; they have significant potential to expand the donor pool. Herein, we analyze the results of transplanted livers and all potential donors treated under our type 2 DCD protocol. Cardiac arrest was witnessed; potential donors arrived at the hospital after attempts at resuscitation had failed. Death was declared based on the absence of cardiorespiratory activity during a 5-min no-touch period. Femoral vessels were cannulated to establish normothermic extracorporeal membrane oxygenation, which was maintained until organ recovery. From April 2002 to December 2010, there were 400 potential donors; 34 liver transplants were performed (9%). Among recipients, median age, model for end-stage liver disease and cold and reperfusion warm ischemic times were 55 years (49-60), 19 (14-21) and 380 (325-430) and 30 min (26-35), respectively. Overall, 236 (59%) and 130 (32%) livers were turned down due to absolute and relative contraindications to donate, respectively. One-year recipient and graft survivals were 82% and 70%, respectively (median follow-up 24 months). The applicability of type 2 DCD liver transplant was <10%; however, with better preservation technology and expanded transplant criteria, we may be able to improve this figure significantly.

Outcome of HCV/HIV-coinfected liver transplant recipients: a prospective and multicenter cohort study.

Eighty-four HCV/HIV-coinfected and 252-matched HCV-monoinfected liver transplant recipients were included in a prospective multicenter study. Thirty-six (43%) HCV/HIV-coinfected and 75 (30%) HCV-monoinfected patients died, with a survival rate at 5 years of 54% (95% CI, 42-64) and 71% (95% CI, 66 to 77; p = 0.008), respectively. When both groups were considered together, HIV infection was an independent predictor of mortality (HR, 2.202; 95% CI, 1.420-3.413 [p < 0.001]). Multivariate analysis of only the HCV/HIV-coinfected recipients, revealed HCV genotype 1 (HR, 2.98; 95% CI, 1.32-6.76), donor risk index (HR, 9.48; 95% CI, 2.75-32.73) and negative plasma HCV RNA (HR, 0.14; 95% CI, 0.03-0.62) to be associated with mortality. When this analysis was restricted to pretransplant variables, we identified three independent factors (HCV genotype 1, pretransplant MELD score and centers with <1 liver transplantation/year in HIV-infected patients) that allowed us to identify a subset of 60 (71%) patients with a similar 5-year prognosis (69%[95% CI, 54-80]) to that of HCV-monoinfected recipients. In conclusion, 5-year survival in HCV/HIV-coinfected liver recipients was lower than in HCV-monoinfected recipients, although an important subset with a favorable prognosis was identified in the former.

Liver retransplantation in HIV-infected patients: a prospective cohort study.

Information regarding liver retransplantation in HIV-infected patients is scant. Data from 14 HIV-infected patients retransplanted between 2002 and 2011 in Spain (6% retransplantation rate) were analyzed and compared with those from 157 matched HIV-negative retransplanted patients. In HIV-infected patients, early (≤30 days) retransplantation was more frequently indicated (57% vs. 29%; p = 0.057), and retransplantation for HCV recurrence was less frequently indicated (7% vs. 37%; p = 0.036). Survival probability after retransplantation in HIV-positive patients was lower than in HIV-negative patients, 42% versus 64% at 3 years, although not significantly (p = 0.160). Among HIV-infected patients, those with undetectable HCV RNA at retransplantation and those with late (>30 days) retransplantation showed better 3-year survival probability (80% and 67%, respectively), similar to that in their respective HIV-negative counterparts (72% and 70%). In HIV-infected and HIV-negative patients, 3-year survival probability in those with positive HCV RNA at retransplantation was 22% versus 65% (p = 0.008); in those with early retransplantation, 3-year survival probability was 25% versus 56% (p = 0.282). HIV infection was controlled with antiretroviral therapy after retransplantation. In conclusion, HIV-infected patients taken as a whole have unsatisfactory survival after liver retransplantation, although patients with undetectable HCV RNA at retransplantation or undergoing late retransplantation show a more favorable outcome.

Comparison of transcriptional and blood cell-phenotypic markers between operationally tolerant liver and kidney recipients.

A proportion of transplant recipients can spontaneously accept their grafts in the absence of immunosuppression (operational tolerance). Previous studies identified blood transcriptional and cell-phenotypic markers characteristic of either liver or kidney tolerant recipients. However, the small number of patients analyzed and the use of different transcriptional platforms hampered data interpretation. In this study we directly compared samples from kidney and liver tolerant recipients in order to identify potential similarities in immune-related parameters. Liver and kidney tolerant recipients differed in blood expression and B-cell immunophenotypic patterns and no significant overlaps were detectable between them. Whereas some recipients coincided in specific NK-related transcripts, this observation was not reproducible in all cohorts analyzed. Our results reveal that certain immune features, but not others, are consistently present across all cohorts of operationally tolerant recipients. This provides a set of reproducible biomarkers that should be explored in future large-scale immunomonitoring trials.

Matrix metalloproteinase 2 in reduced-size liver transplantation: beyond the matrix.

We studied the contribution of matrix metalloproteinase 2 (MMP2) and matrix metalloproteinase 9 (MMP9) to the beneficial effects of preconditioning (PC) in reduced-size orthotopic liver transplantation (ROLT). We also examined the role of c-Jun N-terminal kinase (JNK) and whether it regulates MMP2 in these conditions. Animals were subjected to ROLT with or without PC and pharmacological modulation, and liver tissue samples were then analyzed. We found that MMP2, but notMMP9, is involved in the beneficial effects of PC in ROLT. MMP2 reduced hepatic injury and enhanced liver regeneration. Moreover, inhibition of MMP2 in PC reduced animal survival after transplantation. JNK inhibition in the PC group decreased hepatic injury and enhanced liver regeneration. Furthermore, JNK upregulated MMP2 in PC. In addition, we showed that Tissue inhibitors of matrix metalloproteinases 2 (TIMP2) was also upregulated in PC and that JNK modulation also altered its levels in ROLT and PC. Our results open up new possibilities for therapeutic treatments to reduce I/R injury and increase liver regeneration after ROLT, which are the main limitations in living-donor transplantation.

ATG-Fresenius treatment and low-dose tacrolimus: results of a randomized controlled trial in liver transplantation.

We report the results of a prospective randomized controlled trial in liver transplantation assessing the efficacy and safety of antithymocyte globulin (ATG-Fresenius) plus tacrolimus monotherapy at gradually decreasing doses. Patients were randomized to either: (a) standard-dose tacrolimus plus steroids;or (b) peritransplant ATG-Fresenius plus reduced-dose tacrolimus monotherapy followed by weaning of tacrolimus starting 3 months after transplantation. The primary end-point was the achievement of very low-dose tacrolimus (every-other-day or once daily dose with <5 ng/mL trough levels) at 12 months after transplantation. Acute rejection occurring during the first 3 months after transplantation was more frequent in the ATG group (52.4% vs. 25%). Moreover, late acute rejection episodes occurred in all recipients in whom weaning was attempted and no recipients reached the primary end-point. This motivated the premature termination of the trial. Tacrolimus trough levels were lower in the ATG-Fresenius group but no benefits in terms of improved renal function, lower metabolic complications or increased prevalence of tolerance-related biomarkers were observed. In conclusion, the use of ATG-Fresenius and tacrolimus at gradually decreasing doses was associated with a high rate of rejection, did not allow for the administration of very low doses of tacrolimus and failed to provide detectable clinical benefits. ClinicalTrials.gov identifier: NCT00436722.

Therapeutic targets in liver transplantation: angiotensin II in nonsteatotic grafts and angiotensin-(1-7) in steatotic grafts.

Numerous steatotic livers are discarded as unsuitable for transplantation because of their poor tolerance of ischemia-reperfusion(I/R). The injurious effects of angiotensin (Ang)-II and the benefits of Ang-(1-7) in various pathologies are well documented. We examined the generation of Ang II and Ang-(1-7) in steatotic and nonsteatotic liver grafts from Zucker rats following transplantation. We also studied in both liver grafts the effects of Ang-II receptors antagonists and Ang-(1-7) receptor antagonists on hepatic I/R damage associated with transplantation. Nonsteatotic grafts showed higher Ang II levels than steatotic grafts, whereas steatotic grafts showed higher Ang-(1-7) levels than nonsteatotic grafts. Ang II receptor antagonists protected only nonsteatotic grafts against damage, whereas Ang-(1-7) receptor antagonists were effective only in steatotic grafts. The protection conferred by Ang II receptor antagonists in nonsteatotic grafts was associated with ERK 1/2 overexpression, whereas the beneficial effects of Ang-(1-7) receptor antagonists in steatotic grafts may be mediated by NO inhibition. Our results show that Ang II receptor antagonists are effective only in nonsteatotic liver transplantation and point to a novel therapeutic target in liver transplantation based on Ang-(1-7), which is specific for steatotic liver grafts.

Delayed introduction of reduced-dose tacrolimus, and renal function in liver transplantation: the 'ReSpECT' study.

We report a multicenter, prospective, randomized, open-label trial investigating the effect of lower levels and delayed introduction of tacrolimus on renal function in liver transplant recipients. Adult patients with good renal function undergoing primary liver transplant were randomized to either: group A (standard-dose tacrolimus [target trough levels >10 ng/mL] and corticosteroids; n = 183); group B (mycophenolate mofetil [MMF] 2g/day, reduced-dose tacrolimus [target trough levels

Is the peptide bond formation activated by Cu(2+) interactions? Insights from density functional calculations.

The catalytic role that Cu(2+) cations play in the peptide bond formation has been addressed by means of density functional calculations. First, the Cu(2+)-(glycine)2 --> Cu(2+)-(glycylglycine) + H2O reaction was investigated since mass spectrometry low collision activated dissociation (CAD) spectra of Cu(2+)-(glycine)2 led to the elimination of a water molecule, which suggested that an intracomplex peptide bond formation might have occurred. Results show that this intracomplex condensation is associated to a very high free energy barrier (97 kcal mol(-1)) and reaction free energy (66 kcal mol(-1)) because of the loss of metal coordination during the reaction. Second, on the basis of the salt-induced peptide formation theory, the condensation reaction between two glycines was studied in aqueous solution using discrete water molecules and the conductor polarized continuum model (CPCM) continuous method. It is found that the synergy between the interaction of glycines with Cu(2+) and the presence of water molecules acting as proton-transfer helpers significantly lower the activation barrier (from 55 kcal/mol for the uncatalyzed system to 20 kcal/mol for the Cu(2+) solvated system) which largely favors the formation of the peptide bond.