High-throughput virtual search of small molecules for controlling the mechanical stability of human CD4.

Protein mechanical stability determines the function of a myriad of proteins, especially proteins from the extracellular matrix. Failure to maintain protein mechanical stability may result in diseases and disorders such as cancer, cardiomyopathies, or muscular dystrophy. Thus, developing mutation-free approaches to enhance and control the mechanical stability of proteins using pharmacology-based methods may have important implications in drug development and discovery. Here, we present the first approach that employs computational high-throughput virtual screening and molecular docking to search for small molecules in chemical libraries that function as mechano-regulators of the stability of human cluster of differentiation 4, receptor of HIV-1. Using single-molecule force spectroscopy, we prove that these small molecules can increase the mechanical stability of CD4D1D2 domains over 4-fold in addition to modifying the mechanical unfolding pathways. Our experiments demonstrate that chemical libraries are a source of mechanoactive molecules and that drug discovery approaches provide the foundation of a new type of molecular function, that is, mechano-regulation, paving the way toward mechanopharmacology.

Basal oxidation of conserved cysteines modulates cardiac titin stiffness and dynamics.

Titin, as the main protein responsible for the passive stiffness of the sarcomere, plays a key role in diastolic function and is a determinant factor in the etiology of heart disease. Titin stiffness depends on unfolding and folding transitions of immunoglobulin-like (Ig) domains of the I-band, and recent studies have shown that oxidative modifications of cryptic cysteines belonging to these Ig domains modulate their mechanical properties in vitro. However, the relevance of this mode of titin mechanical modulation in vivo remains largely unknown. Here, we describe the high evolutionary conservation of titin mechanical cysteines and show that they are remarkably oxidized in murine cardiac tissue. Mass spectrometry analyses indicate a similar landscape of basal oxidation in murine and human myocardium. Monte Carlo simulations illustrate how disulfides and S-thiolations on these cysteines increase the dynamics of the protein at physiological forces, while enabling load- and isoform-dependent regulation of titin stiffness. Our results demonstrate the role of conserved cysteines in the modulation of titin mechanical properties in vivo and point to potential redox-based pathomechanisms in heart disease.

Prenatal Diagnosis of Fetal Lymphangioma: A Case Series.

Fetal lymphangioma is an uncommon congenital malformation that is mainly comprised of the subcutaneous tissue of the neck. This malformation can develop in other areas like the thoracic and axillary regions, though rarely. We report 6 consecutive cases of lymphatic malformation in a fetal center in Dominican Republic. In our case series fetal chest lymphangiomas were present in 2 fetuses. In addition, 2 cases of axillary lymphangiomas also involved the thoracic region. Adequate management by a multidiciplinary team is necessary to provide a better approach to delivery.

Fibrosis pulmonar idiopática. Características clínicas de un registro en Panamá. 2017-2020

Introducción: La fibrosis pulmonar idiopática es una enfermedad rara, asociada al envejecimiento, con una alta tasa de mortalidad entre los 2.5 a 3 años. El diagnóstico se realiza con la sospecha clínica y la confirmación del patrón de neumonía intersticial usual, en tomografía del tórax o en biopsia pulmonar; el tratamiento actual se basa en fármacos anti fibróticos. Material y método: Se llevó a cabo un estudio descriptivo y prospectivo tipo registro, de pacientes con fibrosis pulmonar idiopática, que estuviesen en seguimiento por médicos especialistas en neumología en 4 consultorios privados de la ciudad de Panamá entre el 2017 y el 2020. Resultado: Se evaluaron 32 pacientes, edad media de 69 años (rango 53 ­ 91); 18/32 (56.3%) fueron masculinos, y de estos, 16/18 (88.9%) eran fumadores. Los crepitantes tipo velcro subescapulares se encontraron en 29/32 (90.6%). El patrón de neumonía intersticial usual en tomografía se encontró en 30/32 (93.7%), y 2/32 (6.3%) en biopsia. Los medicamentos anti fibróticos fueron usados en 6/32 (18.6%); el 50% usó nintedanib y el 50% pirfenidona. Fallecieron 4/32 (12.5%) pacientes en el periodo de estudio. Conclusión: La fibrosis pulmonar idiopática es frecuente en mayores de 50 años, principalmente varones, con historial de tabaquismo y reflujo gastroesofágico. Los crepitantes tipo velcro subescapulares son un hallazgo frecuente. El diagnóstico se realizó en su mayoría con tomografía de tórax de alta resolución; la difusión de monóxido de carbono y la saturación en la caminata de 6 minutos son las pruebas funcionales más alteradas. Menos de un tercio de los pacientes recibe tratamiento con anti fibróticos. (provisto por Infomedic International)

Introduction: Idiopathic pulmonary fibrosis is a rare disease, associated with aging, with a high mortality rate between 2.5 to 3 years. Diagnosis is made with clinical suspicion and confirmation of the usual interstitial pneumonia pattern in chest tomography or lung biopsy; current treatment is based on antifibrotic drugs. Methods: A descriptive and prospective registry-type study was carried out on patients with idiopathic pulmonary fibrosis who were being followed up by pulmonology specialists in 4 private practices in Panama City between 2017 and 2020. Results: 32 patients were evaluated, mean age 69 years (range 53 - 91); 18/32 (56.3%) were male, and of these, 16/18 (88.9%) were smokers. Subscapularis velcro-type crackles were found in 29/32 (90.6%). The usual interstitial pneumonia pattern on CT was found in 30/32 (93.7%), and 2/32 (6.3%) on biopsy. Antifibrotic drugs were used in 6/32 (18.6%); 50% used nintedanib and 50% pirfenidone. Four/32 (12.5%) patients died during the study period. Conclusion: Idiopathic pulmonary fibrosis is frequent in patients older than 50 years, mainly males, with a history of smoking and gastroesophageal reflux. Subscapular Velcro-like crackles are a frequent finding. Diagnosis was mostly made with high-resolution chest CT; carbon monoxide diffusion and 6-minute walk saturation are the most altered functional tests. Less than one third of patients receive treatment with antifibrotic drugs. (provided by Infomedic International)

Manifestaciones trombóticas en la neumonía viral por SARS-CoV-2 pacientes hospitalizados en el Hospital «Irma de Lourdes Tzanetatos¼, Panamá, Panamá. De marzo de 2020 a septiembre de 2020 / Thrombotic manifestations in viral neumonia by CoV-2-SARS hospitalized patients at «Irma Lourdes Tzanetatos¼ Hospital, Panama, Panama. March 2020 to September 2020

La respuesta a la infección viral produce un estado de trombosis o hipercoagulabilidad que, unido a la inflamación de las células endoteliales, puede generar disfunción plaquetaria y predisposición a la formación de trombos que, aunque con frecuencia son más venosos, también pueden aparecer en el sistema arterial y producir infartos a cualquier nivel así como tromboembolia e hipertensión pulmonar. Estas manifestaciones han sido captadas hospitalariamente y al egreso de los pacientes detectados por SARS-CoV-2 habiendo ya cumplido el tiempo establecido de virulencia. Los criterios diagnósticos de respuesta inmunológica trombótica asociada a COVID-19 (RITAC) ayudan a seleccionar al paciente que está predispuesto a esta condición; a esto se añade que el paciente ya tiene un diagnóstico de infección por SARS-CoV-2 (AU)

The response to viral infection produces a prothrombotic state of hypercoagulability , united with an inflammation of endothelial cells, It can generate platelet dysfunction and predisposition to the formation of thrombus, that, although, are more frequently venous, Also, it can appear in the arterial system and cause heart attacks at any level; thromboembolism and pulmonary hypertension, as well. These manifestations have been captured hospitably and with the egress of patients detected by SARS-CoV-2. The diagnostic criteria of RITAC (abbreviation in Spanish of Thrombotic Immune Response Associated to COVID-19), help to select the patient who is predisposed to this condition; adding that the patient already has a diagnosis of SARS-CoV-2 infection (AU)

CDCA7 finely tunes cytoskeleton dynamics to promote lymphoma migration and invasion.

Metastases, the major cause of death from cancer, require cells' acquisition of the ability to migrate and involve multiple steps, including local tumor cell invasion and basement membrane penetration. Certain lymphoid tumors are highly metastatic, but the mechanisms of invasion by lymphoma cells are poorly understood. We recently showed that CDCA7, a protein induced by MYC, is overexpressed in lymphoid tumors and that its knockdown decreases lymphoid tumor growth without inhibiting the proliferation of normal cells. Here we show that CDCA7 is critical for invasion and migration of lymphoma cells. Indeed, CDCA7 knockdown in lymphoma cells limited tumor cell invasion in matrigel-coated transwell plates and tumor invasion of neighboring tissues in a mouse xenograft model and in a zebrafish model of cell invasion. CDCA7 silencing markedly inhibited lymphoma cell migration on fibronectin without modifying cell adhesion to this protein. Instead, CDCA7 knockdown markedly disrupted the precise dynamic reorganization of actomyosin and tubulin cytoskeletons required for efficient migration. In particular, CDCA7 silencing impaired tubulin and actomyosin cytoskeleton polarization, increased filamentous actin formation, and induced myosin activation. Of note, inhibitors of actin polymerization, myosin II, or ROCK reestablished the migration capacity of CDCA7-silenced lymphoma cells. Given the critical role of CDCA7 in lymphoma-genesis and invasion, therapies aimed at inhibiting its expression or activity might provide significant control of lymphoma growth, invasion, and metastatic dissemination.

Instrumental Effects in the Dynamics of an Ultrafast Folding Protein under Mechanical Force.

The analysis and interpretation of single molecule force spectroscopy (smFS) experiments is often complicated by hidden effects from the measuring device. Here we investigate these effects in our recent smFS experiments on the ultrafast folding protein gpW, which has been previously shown to fold without crossing a free energy barrier in the absence of force (i.e., downhill folding). Using atomic force microscopy (AFM) smFS experiments, we found that a very small force of ∼5 pN brings gpW near its unfolding midpoint and results in two-state (un)folding patterns that indicate the emergence of a force-induced free energy barrier. The change in the folding regime is concomitant with a 30,000-fold slowdown of the folding and unfolding times, from a few microseconds that it takes gpW to (un)fold at the midpoint temperature to seconds in the AFM. These results are puzzling because the barrier induced by force in the folding free energy landscape of gpW is far too small to account for such a difference in time scales. Here we use recently developed theoretical methods to resolve the origin of the strikingly slow dynamics of gpW under mechanical force. We find that, while the AFM experiments correctly capture the equilibrium distance distribution, the measured dynamics are entirely controlled by the response of the cantilever and polyprotein linker, which is much slower than the protein conformational dynamics. This interpretation is likely applicable to the folding of other small biomolecules in smFS experiments, and becomes particularly important in the case of systems with fast folding dynamics and small free energy barriers, and for instruments with slow response times.

The life of proteins under mechanical force.

Although much of our understanding of protein folding comes from studies of isolated protein domains in bulk, in the cellular environment the intervention of external molecular machines is essential during the protein life cycle. During the past decade single molecule force spectroscopy techniques have been extremely useful to deepen our understanding of these interventional molecular processes, as they allow for monitoring and manipulating mechanochemical events in individual protein molecules. Here, we review some of the critical steps in the protein life cycle, starting with the biosynthesis of the nascent polypeptide chain in the ribosome, continuing with the folding supported by chaperones and the translocation into different cell compartments, and ending with proteolysis in the proteasome. Along these steps, proteins experience molecular forces often combined with chemical transformations, affecting their folding and structure, which are measured or mimicked in the laboratory by the application of force with a single molecule apparatus. These mechanochemical reactions can potentially be used as targets for fighting against diseases. Inspired by these insightful experiments, we devise an outlook on the emerging field of mechanopharmacology, which reflects an alternative paradigm for drug design.

The influence of disulfide bonds on the mechanical stability of proteins is context dependent.

Disulfide bonds play a crucial role in proteins, modulating their stability and constraining their conformational dynamics. A particularly important case is that of proteins that need to withstand forces arising from their normal biological function and that are often disulfide bonded. However, the influence of disulfides on the overall mechanical stability of proteins is poorly understood. Here, we used single-molecule force spectroscopy (smFS) to study the role of disulfide bonds in different mechanical proteins in terms of their unfolding forces. For this purpose, we chose the pilus protein FimG from Gram-negative bacteria and a disulfide-bonded variant of the I91 human cardiac titin polyprotein. Our results show that disulfide bonds can alter the mechanical stability of proteins in different ways depending on the properties of the system. Specifically, disulfide-bonded FimG undergoes a 30% increase in its mechanical stability compared with its reduced counterpart, whereas the unfolding force of I91 domains experiences a decrease of 15% relative to the WT form. Using a coarse-grained simulation model, we rationalized that the increase in mechanical stability of FimG is due to a shift in the mechanical unfolding pathway. The simple topology-based explanation suggests a neutral effect in the case of titin. In summary, our results indicate that disulfide bonds in proteins act in a context-dependent manner rather than simply as mechanical lockers, underscoring the importance of considering disulfide bonds both computationally and experimentally when studying the mechanical properties of proteins.

Portal hydatid with secondary cavernomatosis.

The hydatid cyst is a parasitic infection included within the category of zoonoses, in which there exists a direct or indirect relation with animals, particularly with dogs. We report a clinical case of a patient who has undergone surgery of hydatid cyst in the right hepatic lobe. Seven years later the patient presents hydatid cyst in portal vein with secondary cavernomatosis, which is a rare complication. There are few cases described in the literature. The symtomatology presented by hydatid cyst is variable and the diagnosis is made by ultrasonography, TC and/or RNM. The treatment of choice is the surgical removal of the cyst, prior to surgery Albendazol must be administered for 1-2 weeks and be maintained for 4 more weeks after surgery.

Influencia de las clases de educación física sobre el nivel de actividad física medido a través de una aplicación móvil en adolescentes / Influence of physical education classes on the level of physical activity measured through a mobile application in adolescents

La literatura científica ha demostrado a través de diversos métodos (cuestionarios y acelerómetros, principalmente) que las clases de educación física (EF) influyen positivamente en el logro de las recomendaciones de actividad física (AF) en la población escolar. Sin embargo, el uso de nuevas tecnologías como los teléfonos móviles o smartphone, parecen ser una interesante herramienta de bajo costo que también podrían ayudar a determinar el nivel de AF. Por tanto, el objetivo del presente estudio fue describir como influye la clase de EF en el número de pasos diarios en escolares que utilizaron una aplicación móvil “RuntasticPedometer” para monitorear dicha variable. La muestra estuvo compuesta por 50 estudiantes (edad media de 14.1 +/-1.6 años) pertenecientes al Instituto de Educación Secundaria “Melchor de Macanaz” de la localidad de Hellín (Albacete, España). Tras analizar los datos, se observaron diferencias significativas (p<0.05) durante los días en que los escolares realizaron la clase de EF y los que no (9.950 vs. 8.023 pasos/día,respectivamente). Además, la variación entre los días con y sin EF también fueron estadísticamente diferentes entre niños y niñas (2.644 vs. 1.579 pasos/día, respetivamente).Finalmente, se podría concluir que las clases de EF parecen influir positivamente en el aumento del número de pasos en escolares, lo que elevaría las posibilidades de que estos niños y niñas logren los parámetros mínimos de AF recomendada por las diversas organizaciones de salud. Por otro lado, cabe señalar que el uso de nuevas tecnologías de uso masivo como los smartphone, podrían ser una interesante herramienta a validar orientadas al monitoreo de la AF.

The scientific literature has demonstrated through various methods (questionnaires and accelerometers, mainly), the classes of physical education (PE) positively influence the achievement of the recommendations of physical activity (PA) in the school population. However, the use of new technologies such as mobile phones or smartphone, appear to be an interesting inexpensive tool that could also help determine the level of AF. Therefore, the objective of this study was to describe as PE class influences in the daily number of steps in school who used a mobile application "Runtastic Pedometer" to monitor the variable. The sample consisted of 50 students (mean age 14.1 +/- 1.6 years) belonging to the Institute of Secondary Education "MelchorMacanaz" from the Hellín (Albacete, Spain). After analyzing the data, significant differences (p <0.05) were observed during the days when the school made PE class and those without (8,023 vs. 9,950 steps / day, respectively). Moreover, the variations between days with and without EF were also statistically different between boys and girls (2,644 vs. 1,579 steps/day,respectively). Finally, one could conclude that PE classes seem positively influence the increase in the number of steps in school raising the chances that these children achieve the minimum standards recommended HF various health organizations. On the other hand, it notes that the use of new technologies in widespread use as the smartphone could be an interesting tool to validate oriented monitoring AF.

Prolactin modulates cytokine production induced by culture filtrate proteins of M. bovis through different signaling mechanisms in THP1 cells.

The immunomodulatory functions of prolactin (PRL) are well recognized. Augmented PRL plasma levels were observed in patients with advanced tuberculosis (TB). Recently, we have reported that LPS and Mycobacterium bovis (M. bovis) induced differential expression of PRL receptor (PRLR) isoforms in THP-1 cells and bovine macrophages, respectively. The aim of this work was to determine whether PRL should be considered as a potential modulator of the signaling pathways and cytokine synthesis, induced by culture filtrate protein (CFP) from M. bovis in THP-1 monocytes. The THP-1 cells were stimulated with PRL (20ng/mL), M. bovis CFP (50µg/mL). PRLR as well as phosphorylated STAT3, STAT5, Akt1/2/3, ERK1/2 and p38 expression were evaluated by Western blot. IL1-ß, TNF-α, IL-6, IL-12, IL-8, and IL-10 concentrations were measured by ELISA. Our results demonstrated that the expression pattern of PRLR short isoforms is induced by M. bovis CFP. M bovis CFP induced phosphorylation of Akt2, ERK1/2, p38, STAT3, and STAT5 pathways. In turn, PRL only activated the JAK2/STAT3-5 signaling pathway. However, when combined both stimuli, PRL significantly increased STAT3-5 phosphorylation and downregulated Akt2, ERK1/2, and p38 phosphorylation. As expected, M. bovis CFP induced substantial amounts of IL1-ß, IL-6, TNF-α, IL-8, IL-12, and IL-10. However, the PRL costimulation considerably decreased IL1-ß, TNF-α, and IL-12 secretion, and increased IL-10 production. This results suggest that up-regulation of IL-10 by PRL might be modulating the pro-inflammatory response against mycobacterial antigens through the MAPK pathway.

Estado y supervivencia de dientes monorradiculares con formadentinas elaborados en facultad de odontología de universidad de el salvador en período 2010-2012 / Status and survival of monoradicular teeth with formadentines elaborated in the dental school of the university of el salvador in the period 2010-2012

Objetivo: Determinar el estado y supervivencia de dientes monorradiculares con formadentinas elaborados en Facultad de Odontología de Universidad de El Salvador en período 2010-2012. Metodología: La investigación incluyó una muestra de 126 dientes monorradiculares tratados con formadentina, de pacientes atendidos en la Facultad de Odontología durante el período 2010-2012. Se realizó evaluación clínica y radiográfica, registrándose los datos de cada paciente en una guía de observación en donde se obtuvieron resultados de supervivencia del diente tratado con formadentina, presencia o ausencia de dolor, movilidad, entre otros hallazgos. Los datos se vaciaron en el programa SPSS v.19. Resultados: El 88.08% de los casos registró una supervivencia igual o mayor a 2 años 6 meses. Respecto al estado clínico el 65.9% presentó corona definitiva como restauración, el 72.3% no presentó dolor a la percusión y el 76.5% ausencia de movilidad dental. Al estado radiográfico se encontró que el 73.9% no presentó ensanchamiento del ligamento periodontal, el 65.5% no evidenció lesiones periapicales, mientras que el 98.3% no reflejó reabsorciones internas y el 95% no mostró reabsorciones externas; el 99.2% no presentó fracturas dentales. Conclusiones: La supervivencia de los dientes tratados con formadentina fue igual o mayor a 2 años 6 meses, el estado clínico y radiográfico de los dientes con formadentina fue bueno ya que la mayoría de los dientes presentaron características ideales al momento de la evaluación. Palabras claves: dientes monorradiculares, estado clínico, estado radiográfico, supervivencia, formadentina.

Objective: To determine survival and status of rooted teeth with post cast made in Faculty of Dentistry, University of El Salvador in 2010-2012 Methodology: The investigation includes a sample of 126 monoradicular teeth restored with cast post in patients attended on the Faculty of Dentistry in the period 2010-2012. They were evaluated clinically and radiographically recording the data in an observational guide where we obtained results of survival of the teeth with post cast, presence or absence of pain, mobility and other findings. The data was cleared in the program SPSS v. 19. Results: The 88.08% of the cases showed a survival equal to or greater than 2 years 6 months. Regarding clinical status the 65.9% presented dental crown as final restoration, also the 72.3% didn´t present pain to percussion and the 76.5% didn´t present any grade of mobility. At the radiographic evaluation it was found that the 73.9% didn´t present widening of the periodontal ligament, the 65.5% didn´t show periapical lesions while the 98.3% didn´t show inner resorption and the 95% didn´t present external resorption; the 99.2% didn´t show teeth fractures. Conclusions: The survival of restored teeth with cast post was equal or greater than 2 years and 6 months, the clinical and radiographic status of the teeth restored with cast post was good because most teeth showed ideal characteristics at the time of evaluation.

[Pancreatic metastases due to renal carcinoma. Our cases and a literature review]. / Metástasis pancreáticas por carcinoma renal. Nuestra casuística y revisión de la literatura.

OBJECTIVE: To analyse the cases of pancreatic metastases due to renal carcinoma operated on in our hospital between the years 2000 and 2011. MATERIAL AND METHODS: A retrospective study using the variables of 8 patients who were subjected to surgery of pancreatic metastases due to renal carcinoma, and a comparison of our data with those from the literature. RESULTS: The incidence of metastatic disease of the pancreas due to renal carcinoma in our series was 1.2%. All the metastases were metachronous, with both sexes being affected equally. The mean time between resection of the renal tumour and the diagnosis of the metastasis was 12.42 years (range: 1.62-30.13 years). The therapeutic approach to the pancreatic lesions was surgical in all cases. Seven patients are currently still alive. CONCLUSION: Metastatic disease of the pancreas due to renal carcinoma is uncommon (1%-2.8%). The interval between the primary resection and the metastasis can be quite long. Pancreatic metastasis must always be suspected in patients who present with a pancreatic mass and a history of renal carcinoma. Aggressive surgical treatment is recommended in selected cases. The surgery in these cases improves survival and the quality of life.

Effect of neoadjuvant chemotherapy in hepatic steatosis.

UNLABELLED: Chemotherapy drugs often produce side effects in the liver. In recent years, there has been speculation about the ability to produce hepatic steatosis in patients treated with 5-fluorouracil and oxaliplatin. This prospective study examines whether these drugs can produce steatosis in patients with neoadjuvant treatment who were operated on for liver tumors. PURPOSE: Our objective was to assess the effect of neoadjuvant chemotherapy (NAC) on the development of hepatic steatosis in the healthy liver. PATIENTS AND METHODS: This was a prospective study based on 32 patients divided into two groups. The presence of steatosis was assessed using a histological score (Kleiner classification) and a biochemical method (Folch method) for patients from both groups. RESULTS: A total of 14 patients (44%) had hepatic steatosis and half of these were in each group. The steatosis was moderate to severe (grades 2-3) in 4 patients (13%), 2 in each group. The mean levels of triglycerides in the liver were 33.38 and 29.94 mg/g in group I and group II, respectively, with the difference not being statistically significant. CONCLUSIONS: Almost half of the patients treated with NAC for liver neoplasia developed steatosis. Nevertheless, NAC does not seem to increase the risk of hepatic steatosis.

Dwell time analysis of a single-molecule mechanochemical reaction.

Force-clamp spectroscopy is a novel technique for studying mechanochemistry at the single-bond level. Single disulfide bond reduction events are accurately detected as stepwise increases in the length of polyproteins that contain disulfide bonds and that are stretched at a constant force with the cantilever of an atomic force microscope (AFM). The kinetics of this reaction has been measured from single-exponential fits to ensemble averages of the reduction events. However, exponential fits are notoriously ambiguous to use in cases of kinetic data showing multiple reaction pathways. Here we introduce a dwell time analysis technique, of widespread use in the single ion channel field, that we apply to the examination of the kinetics of reduction of disulfide bonds measured from single-molecule force-clamp spectroscopy traces. In this technique, exponentially distributed dwell time data is plotted as a histogram with a logarithmic time scale and a square root ordinate. The advantage of logarithmic histograms is that exponentially distributed dwell times appear as well-defined peaks in the distribution, greatly enhancing our ability to detect multiple kinetic pathways. We apply this technique to examine the distribution of dwell times of 4488 single disulfide bond reduction events measured in the presence of two very different kinds of reducing agents: tris-(2-carboxyethyl)phosphine hydrochloride (TCEP) and the enzyme thioredoxin (TRX). A different clamping force is used for each reducing agent to obtain distributions of dwell times on a similar time scale. In the case of TCEP, the logarithmic histogram of dwell times showed a single peak, corresponding to a single reaction mechanism. By contrast, similar experiments done with TRX showed two well-separated peaks, marking two distinct modes of chemical reduction operating simultaneously. These experiments demonstrate that dwell time analysis techniques are a powerful approach to studying chemical reactions at the single-molecule level.