Arsenic exposure and oxidative damage to lipid, DNA, and protein among general Chinese adults: A repeated-measures cross-sectional and longitudinal study.

Arsenic-related oxidative stress and resultant diseases have attracted global concern, while longitudinal studies are scarce. To assess the relationship between arsenic exposure and systemic oxidative damage, we performed two repeated measures among 5236 observations (4067 participants) in the Wuhan-Zhuhai cohort at the baseline and follow-up after 3 years. Urinary total arsenic, biomarkers of DNA oxidative damage (8-hydroxy-2'-deoxyguanosine (8-OHdG)), lipid peroxidation (8-isoprostaglandin F2alpha (8-isoPGF2α)), and protein oxidative damage (protein carbonyls (PCO)) were detected for all observations. Here we used linear mixed models to estimate the cross-sectional and longitudinal associations between arsenic exposure and oxidative damage. Exposure-response curves were constructed by utilizing the generalized additive mixed models with thin plate regressions. After adjusting for potential confounders, arsenic level was significantly and positively related to the levels of global oxidative damage and their annual increased rates in dose-response manners. In cross-sectional analyses, each 1% increase in arsenic level was associated with a 0.406% (95% confidence interval (CI): 0.379% to 0.433%), 0.360% (0.301% to 0.420%), and 0.079% (0.055% to 0.103%) increase in 8-isoPGF2α, 8-OHdG, and PCO, respectively. More importantly, arsenic was further found to be associated with increased annual change rates of 8-isoPGF2α (ß: 0.147; 95% CI: 0.130 to 0.164), 8-OHdG (0.155; 0.118 to 0.192), and PCO (0.050; 0.035 to 0.064) in the longitudinal analyses. Our study suggested that arsenic exposure was not only positively related with global oxidative damage to lipid, DNA, and protein in cross-sectional analyses, but also associated with annual increased rates of these biomarkers in dose-dependent manners.

Lifting the Concentration Limit of Mass Photometry by PEG Nanopatterning.

Mass photometry (MP) is a rapidly growing optical technique for label-free mass measurement of single biomolecules in solution. The underlying measurement principle provides numerous advantages over ensemble-based methods but has been limited to low analyte concentrations due to the need to uniquely and accurately quantify the binding of individual molecules to the measurement surface, which results in diffraction-limited spots. Here, we combine nanoparticle lithography with surface PEGylation to substantially lower surface binding, resulting in a 2 orders of magnitude improvement in the upper concentration limit associated with mass photometry. We demonstrate the facile tunability of degree of passivation, enabling measurements at increased analyte concentrations. These advances provide access to protein-protein interactions in the high nanomolar to low micromolar range, substantially expanding the application space of mass photometry.

Sour orange (Citrus aurantium) seed, a rich source of protein isolate and hydrolysate - A thorough investigation.

Sour orange (Citrus aurantium) seeds are typically discarded by juice processors as waste. This study aimed to extract protein isolates, produce hydrolysates from de-oiled sour orange seeds (SOS), and characterize their physicochemical properties. Previous studies have described methods to obtain protein isolates and hydrolysates from agricultural residues. However, there is limited data on the SOS. This research characterized protein isolates and hydrolysates from SOS, emphasizing yield, purity, and amino acid composition. Protein isolates were extracted using borate saline buffer, saline, and distilled water. Enzymatically hydrolysis was conducted using Protamex® (a commercial protease) at concentrations ranging from 0.2 to 5 g enzyme/100g protein isolate. Differential scanning calorimetry, electrophoresis, and FT-IR spectroscopy were utilized to characterize the isolates and hydrolysates. Data showed that using 5 % saline resulted in protein extraction with a yield and purity of 30 and 86 %, respectively. DSC analysis revealed that the denaturation temperature of the protein isolate was 68 °C, while the hydrolysates exhibited structural instability, as indicated by a decrease in enthalpy change compared to the isolate. The protein isolate had a 76° contact angle. The amino acid profile showed a significant presence of glutamic acid (130.530 mg/g) and arginine (70.210 mg/g). Electrophoresis analysis exhibited four major bands of the protein. The bands' intensity decreased, and new bands appeared after hydrolysis. The enzyme hydrolysis was confirmed using the O-phthaldialdehyde method and FTIR. Findings revealed that based on the free amine group quantity, the hydrolysate obtained using 5 g enzyme/100g protein isolate was 14.220 ± 0.299 µmol/mg protein. The study concluded that sour orange seeds are a good source of protein, with protein isolates and hydrolysates exhibiting desirable characteristics. More research needs to be conducted to acquire further information about their functional properties and potential applications.

Exploring the causal relationship between interleukin-6 or C reactive protein and malignant melanoma using a two-sample Mendelian randomization approach.

The goal was to explore the effect of interleukin-6 (IL6) and C reactive protein (CRP) on malignant melanoma (MM) using two-sample Mendelian randomization. Methods: Data for this study were obtained from the IEU Open GWAS project website for genome-wide association study data (GWAS) on interleukin-6, C reactive protein levels and malignant melanoma. Inverse variance weighted (IVW) method was mainly used and supplemented with MR-Egger regression and weighted median. Finally, horizontal multivariate validity and heterogeneity tests were performed to assess the stability and reliability of the results. Results: The results of univariate two-sample MR analyses showed no significant effect of CRP on MM: inverse variance weighting method (OR=0.999, 95% CI: 0.998-1.001, P=0.343), MR-Egger regression (OR= 1.000, 95% CI: 0.998-1.001, P= 0.180), and weighted median method (OR= 0.999, 95% CI: 0.997 to 1.000, P= 0.583), and weighted model (OR= 0.999, 95% CI: 0.998 to 1.001, P= 0.328). Also,IL-6 had no significant effect on MM: inverse variance weighting method (OR= 1.001, 95% CI: 0.999 to 1.002, P=0.461), MR-Egger regression (OR= 1.000, 95% CI: 0.997 to 1.004, P= 0.910), weighted median method (OR= 1.000, 95% CI: 0.998 to 1.002, P= 0.749), and weighted mode (OR= 1.000, 95% CI: 0.998 to 1.002, P= 0.820). Conclusion: There was no causal relationship between C-reactive protein and IL-6 on the risk of malignant melanoma.

CREB3 suppresses hepatocellular carcinoma progression by depressing AKT signaling through competitively binding with insulin receptor and transcriptionally activating RNA-binding motif protein 38.

cAMP responsive element binding protein 3 (CREB3), belonging to bZIP family, was reported to play multiple roles in various cancers, but its role in hepatocellular carcinoma (HCC) is still unclear. cAMP responsive element binding protein 3 like 3 (CREB3L3), another member of bZIP family, was thought to be transcription factor (TF) to regulate hepatic metabolism. Nevertheless, except for being TFs, other function of bZIP family were poorly understood. In this study, we found CREB3 inhibited growth and metastasis of HCC in vitro and in vivo. RNA sequencing indicated CREB3 regulated AKT signaling to influence HCC progression. Mass spectrometry analysis revealed CREB3 interacted with insulin receptor (INSR). Mechanistically, CREB3 suppressed AKT phosphorylation by inhibiting the interaction of INSR with insulin receptor substrate 1 (IRS1). In our study, CREB3 was firstly proved to affect activation of substrates by interacting with tyrosine kinase receptor. Besides, CREB3 could act as a TF to transactivate RNA-binding motif protein 38 (RBM38) expression, leading to suppressed AKT phosphorylation. Rescue experiments further confirmed the independence between the two functional manners. In conclusion, CREB3 acted as a tumor suppressor in HCC, which inhibited AKT phosphorylation through independently interfering interaction of INSR with IRS1, and transcriptionally activating RBM38.

Aloe vera supplementation improves cardiovascular risk factors in hemodialysis patients: A randomized, double-blind, placebo-controlled trial.

Objective: This study assessed the effects of Aloe vera supplementation on serum inflammatory factors, blood sugar and lipid profiles in hemodialysis patients. Materials and Methods: Totally, 50 hemodialysis patients were allocated randomly to either Aloe vera or placebo groups. The Aloe vera group received 2 Aloe vera capsules daily for 8 weeks (500 mg/day). Serum C-reactive protein (hs- CRP), Fasting blood glucose (FBS), and lipid profiles levels were evaluated at the baseline and the end of the eighth week. Results: Aloe vera supplementation for 8 weeks was associated with a significant reduction of serum hs- CRP (p=0.004), total cholesterol (p=0.01), low density lipoprotein (LDL) (p=0.02) leves and increased high density lipoprotein (HDL) (p=0.002) concentration in the hemodialysis patients. Conclusion: Aloe vera supplementation is beneficial in improvement of cardiovascular risk factors in hemodialysis patients.

The first report of single nucleotide polymorphisms in the open reading frame of the prion-like protein gene in rabbits.

Background: Natural cases of prion disease have not been reported in rabbits, and prior attempts to identify a prion conversion agent have been unsuccessful. However, recent applications of prion seed amplifying experimental techniques have sparked renewed interest in the potential susceptibility of rabbits to prion disease infections. Among several factors related to prion disease, polymorphisms within the prion-like protein gene (PRND), a member of the prion protein family, have been reported as significantly associated with disease susceptibility in various species. Therefore, our study aimed to investigate polymorphisms in the PRND gene of rabbits and analyze their genetic characteristics. Methods: Genomic DNA was extracted from 207 rabbit samples to investigate leporine PRND polymorphisms. Subsequently, amplicon sequencing targeting the coding region of the leporine PRND gene was conducted. Additionally, linkage disequilibrium (LD) analysis was employed to assess the connection within and between loci. The impact of non-synonymous single nucleotide polymorphisms (SNPs) on the Doppel protein was evaluated using PolyPhen-2. Results: We found nine novel SNPs in the leporine PRND gene: c.18A > G, c.76G > C, c.128C > T, c.146C > T, c.315A > G, c.488G > A, c.525G > C, c.544G > A, and c.579A > G. Notably, seven of these PRND SNPs, excluding c.525G > C and c.579A > G, exhibited strong LD values exceeding 0.3. In addition, LD analysis confirmed a robust link between PRNP SNP c.234C > T and PRND SNPs at c.525G > C and c.579A > G. Furthermore, according to PolyPhen-2 and SIFT analyses, the four non-synonymous SNPs were predicted to have deleterious effects on the function or structure of the Doppel protein. However, PANTHER and Missense3D did not indicate such effects. Conclusion: In this paper, we have identified novel SNPs in the rabbit PRND gene and predicted their potential detrimental effects on protein function or structure through four non-synonymous SNPs. Additionally, we observed a genetic linkage between SNPs in the PRND and PRNP genes. These findings may provide insights into understanding the characteristics of rabbits as partially resistant species. To the best of our knowledge, this study is the first to genetically characterize PRND SNPs in rabbits.

Coexistence of nonfluorescent chromoproteins and fluorescent proteins in massive Porites spp. corals manifesting a pink pigmentation response.

Introduction: Several fluorescent proteins (FPs) and chromoproteins (CPs) are present in anthozoans and play possible roles in photoprotection. Coral tissues in massive corals often display discoloration accompanied by inflammation. Incidences of the pink pigmentation response (PPR) in massive Porites, described as inflammatory pink lesions of different shapes and sizes, has recently increased worldwide. FPs are reported to be present in PPR lesions, wherein a red fluorescent protein (RFP) appears to play a role in reducing reactive oxygen species. However, to date, the biochemical characterization and possible roles of the pigments involved are poorly understood. The present study aimed to identify and characterize the proteins responsible for pink discoloration in massive Porites colonies displaying PPRs, as well as to assess the differential distribution of pigments and the antioxidant properties of pigmented areas. Method: CPs were extracted from PPR lesions using gel-filtration chromatography and identified via genetic analysis using liquid chromatography-tandem mass spectrometry. The coexistence of CPs and RFP in coral tissues was assessed using microscopic observation. Photosynthetic antivity and hydrogen peroxide-scavenging activitiy were measured to assess coral stress conditions. Results: The present study revealed that the same CP (plut2.m8.16902.m1) isolated from massive Porites was present in both the pink spot and patch morphologies of the PPR. CPs were also found to coexist with RFP in coral tissues that manifested a PPR, with a differential distribution (coenosarc or tip of polyps' tentacles). High hydrogen peroxide-scavenging rates were found in tissues affected by PPR. Discussion and Conclusion: The coexistence of CPs and RFP suggests their possible differential role in coral immunity. CPs, which are specifically expressed in PPR lesions, may serve as an antioxidant in the affected coral tissue. Overall, this study provides new knowledge to our understanding of the role of CPs in coral immunity.

Variable Expression Common Immune Deficiency: A Report of Five Cases.

We present five cases of common variable immunodeficiency (CVID), comprising three women and two men with a mean age of 23.8 ± 9.2 years. All our patients suffered from recurrent bronchopneumonitis, with complications of purulent pleurisy in two cases, requiring decortication in one case, and resulting in bronchiectasis in three cases. Digestive tract infections were observed in four patients, while two patients presented with ENT infections. One case was complicated by bacterial meningitis. All patients presented with global hypogammaglobulinemia, with CVID and granulomatous manifestation in one case. Treatment consisted of monthly immunoglobulin infusions.

Timing of feeding a protein supplement on nitrogen balance and plasma amino acids during exercise recovery in horses.

Eight geldings weighing 544 ± 16 kg were used to evaluate feeding a postexercise protein meal on plasma amino acids during recovery. Horses were fed sweet feed, corn, grass hay and equal amounts of a protein pellet (32% CP) with meals (MP group) or postexercise (EP group). Horses exercised 1-2 h/day, 5 days/week, for 12 weeks. A pre and poststudy 4 days total urine and feces collection was conducted. Urine and fecal samples were analyzed for nitrogen (N) to calculate N balance. Blood samples were drawn immediately after and at 1 and 3 h postexercise at the start and end of the study for plasma amino acid concentrations. Absorbed N and N retention were greater for the MP group compared to the EP group (p = 0.038, 0.033 respectively). An interaction revealed an increase in fecal N (p = 0.01) and decreased N digestibility for the MP group compared to the EP group at the end of the study. Plasma concentrations for 8 out of 14 amino acids were less for the EP group immediately after exercise compared to the MP group (p < 0.02). Plasma concentrations of lysine and arginine were greater for the EP group compared to the MP group at 1 and 3 h after exercise (p < 0.05 and 0.04 respectively). Changes were different for 8 out of the 14 amino acids immediately post exercise, 7 out of 14 amino acids at 1 h postexercise and 10 out of 14 amino acids at 3 h postexercise with positive changes for the EP group and negative changes for the MP group. The EP group had improved supply of plasma amino acids in the recovery period that sustained for 3 h postexercise and are indicative of better amino acid supply supporting muscle development.

Advancing mRNA Therapeutics: The Role and Future of Nanoparticle Delivery Systems.

The coronavirus (COVID-19) pandemic has underscored the critical role of mRNA-based vaccines as powerful, adaptable, readily manufacturable, and safe methodologies for prophylaxis. mRNA-based treatments are emerging as a hopeful avenue for a plethora of conditions, encompassing infectious diseases, cancer, autoimmune diseases, genetic diseases, and rare disorders. Nonetheless, the in vivo delivery of mRNA faces challenges due to its instability, suboptimal delivery, and potential for triggering undesired immune reactions. In this context, the development of effective drug delivery systems, particularly nanoparticles (NPs), is paramount. Tailored with biophysical and chemical properties and susceptible to surface customization, these NPs have demonstrated enhanced mRNA delivery in vivo and led to the approval of several NPs-based formulations for clinical use. Despite these advancements, the necessity for developing a refined, targeted NP delivery system remains imperative. This review comprehensively surveys the biological, translational, and clinical progress in NPs-mediated mRNA therapeutics for both the prevention and treatment of diverse diseases. By addressing critical factors for enhancing existing methodologies, it aims to inform the future development of precise and efficacious mRNA-based therapeutic interventions.

A NAC transcription factor MNAC3-centered regulatory network negatively modulates rice immunity against blast disease.

NAC transcription factors (TFs) are pivotal in plant immunity against diverse pathogens. Here, we report the functional and regulatory network of MNAC3, a novel NAC TF, in rice immunity. MNAC3, a transcriptional activator, negatively modulates rice immunity against blast and bacterial leaf blight diseases and pathogen-associated molecular pattern (PAMP)-triggered immune responses. MNAC3 binds to a CACG cis-element and activates the transcription of immune-negative target genes OsINO80, OsJAZ10, and OsJAZ11. The negative function of MNAC3 in rice immunity depends on its transcription of downstream genes such as OsINO80 and OsJAZ10. MNAC3 interacts with immunity-related OsPP2C41 (a protein phosphatase), ONAC066 (a NAC TF), and OsDjA6 (a DnaJ chaperone). ONAC066 and OsPP2C41 attenuate MNAC3 transcriptional activity, while OsDjA6 promotes it. Phosphorylation of MNAC3 at S163 is critical for its negative functions in rice immunity. OsPP2C41, which plays positive roles in rice blast resistance and chitin-triggered immune responses, dephosphorylates MNAC3, suppressing its transcriptional activity on the target genes OsINO80, OsJAZ10, and OsJAZ11 and promoting the translocation of MNAC3 from nucleus to cytoplasm. These results establish a MNAC3-centered regulatory network in which OsPP2C41 dephosphorylates MNAC3, attenuating its transcriptional activity on downstream immune-negative target genes in rice. Together, these findings deepen our understanding of molecular mechanisms in rice immunity and offer a novel strategy for genetic improvement of rice disease resistance.

Malnutrition management in children with chronic kidney disease.

Chronic kidney disease (CKD) encompasses diverse conditions such as congenital anomalies, glomerulonephritis, and hereditary nephropathies, necessitating individualized nutritional interventions. Early detection is pivotal due to the heightened risk of adverse outcomes, including compromised growth and increased healthcare costs. The nutritional assessment in pediatric CKD employs a comprehensive, multidisciplinary approach, considering disease-specific factors, growth metrics, and dietary habits. The prevalence of malnutrition, as identified through diverse tools and guidelines, underscores the necessity for regular and vigilant monitoring. Nutritional management strategies seek equilibrium in calorie intake, protein requirements, and electrolyte considerations. Maintaining a well-balanced nutritional intake is crucial for preventing systemic complications and preserving the remaining kidney function. The nuanced landscape of enteral nutrition, inclusive of gastrostomy placement, warrants consideration in scenarios requiring prolonged support, with an emphasis on minimizing risks for optimized outcomes. In conclusion, the ongoing challenge of managing nutrition in pediatric CKD necessitates continuous assessment and adaptation. This review underscores the significance of tailored dietary approaches, not only to foster growth and prevent complications but also to enhance the overall quality of life for children grappling with CKD.

Implicating clinical utility of altered expression of PTCH1 & SMO in oral squamous cell carcinoma.

INTRODUCTION: Oral cancer poses a significant burden on public health in India, with higher incidence and mortality rates. Despite advancements in treatment modalities, prognosis remains poor due to factors such as localized recurrence and lymph node metastasis, potentially influenced by cancer stem cells. Among signaling pathways implicated in CSC regulation, the Hedgehog pathway plays a crucial role in oral squamous cell carcinoma (OSCC). MATERIAL & METHODS: 97 OSCC patients' tissue samples were collected and subjected to RNA isolation, cDNA synthesis and quantitative real-time PCR to analyze PTCH1 and SMO expression. Protein expression was assessed through immunohistochemistry. Clinicopathological parameters were correlated with gene and protein expression. Statistical analysis included Pearson chi-square tests, co-relation co-efficient tests, Kaplan-Meier survival analysis and ROC curve analysis. RESULTS: PTCH1 expression correlated with lymphatic permeation (p = 0.002) and tumor stage (p = 0.002), while SMO expression correlated with lymph node status (p = 0.034) and tumor stage (p = 0.021). PTCH1 gene expression correlated with lymph node status (p = 0.024). High PTCH1 gene expression was associated with shorter survival in tongue cancer patients. ROC curve analysis indicated diagnostic potential for PTCH1 and SMO gene and cytoplasmic SMO expression in distinguishing malignant tissues from adjacent normal tissues. CONCLUSION: PTCH1 and SMO play a crucial role in oral cancer progression, correlating with tumor stages and metastatic potential. Despite not directly influencing overall survival, PTCH1 expression at specific anatomical sites hints at its prognostic implications. PTCH1 and SMO exhibit diagnostic potential, suggesting their utility as molecular markers in oral cancer management and therapeutic strategies.

Discovery and Quantitative Dissection of Cytokinesis Mechanisms Using Dictyostelium discoideum.

The social amoeba Dictyostelium discoideum is a versatile model for understanding many different cellular processes involving cell motility including chemotaxis, phagocytosis, and cytokinesis. Cytokinesis, in particular, is a model cell-shaped change process in which a cell separates into two daughter cells. D. discoideum has been used extensively to identify players in cytokinesis and understand how they comprise the mechanosensory and biochemical pathways of cytokinesis. In this chapter, we describe how we use cDNA library complementation with D. discoideum to discover potential regulators of cytokinesis. Once identified, these regulators are further analyzed through live cell imaging, immunofluorescence imaging, fluorescence correlation and cross-correlation spectroscopy, micropipette aspiration, and fluorescence recovery after photobleaching. Collectively, these methods aid in detailing the mechanisms and signaling pathways that comprise cell division.

APEX2-Mediated Proximity Protein Labeling in Dictyostelium.

Largely due to its simplicity, while being more like human cells compared to other experimental models, Dictyostelium continues to be of great use to discover basic molecular mechanisms and signaling pathways underlying evolutionarily conserved biological processes. However, the identification of new protein interactions implicated in signaling pathways can be particularly challenging in Dictyostelium due to its extremely fast signaling kinetics coupled with the dynamic nature of signaling protein interactions. Recently, the proximity labeling method using engineered ascorbic acid peroxidase 2 (APEX2) in mammalian cells was shown to allow the detection of weak and/or transient protein interactions and also to obtain spatial and temporal resolution. Here, we describe a protocol for successfully using the APEX2-proximity labeling method in Dictyostelium. Coupled with the identification of the labeled proteins by mass spectrometry, this method expands Dictyostelium's proteomics toolbox and should be widely useful for identifying interacting partners involved in a variety of biological processes in Dictyostelium.

Analysis of cGMP Signaling in Dictyostelium.

Biochemical assays are described to analyze signal transduction by the second messenger cGMP in Dictyostelium. The methods include enzyme assays to measure the activity and regulation of cGMP synthesizing guanylyl cyclases and cGMP-degrading phosphodiesterases. In addition, several methods are described to quantify cGMP levels. The target of cGMP in Dictyostelium is the large protein GbpC that has multiple domains including a Roc domain, a kinase domain, and a cGMP-stimulated Ras-GEF domain. A cGMP-binding assay is described to detect and quantify GbpC.

Quantitative Monitoring of GPCR-Mediated Spatiotemporal IP3 Dynamics Using Confocal Fluorescence Microscopy.

Activation of G protein-coupled receptors upon chemoattractant stimulation induces activation of multiple signaling pathways. To fully understand how these signaling pathway coordinates to achieve directional migration of neutrophils, it is essential to determine the dynamics of the spatiotemporal activation profile of signaling components at the level of single living cells. Here, we describe a detailed methodology for monitoring and quantitatively analyzing the spatiotemporal dynamics of 1,4,5-inositol trisphosphate (IP3) in neutrophil-like HL60 cells in response to various chemoattractant fields by applying Förster resonance energy transfer (FRET) fluorescence microscopy.

Proximity-Dependent Activation of Split DNAzyme Kinase.

Binary (also known as split) nucleic acid enzymes have emerged as novel tools in biosensors. We report a new split strategy to split the DNAzyme kinase into two independent and non-functional fragments, denoted DK1sub and DK1enz. In the presence of the specific target, their free ends are brought sufficiently close to interact with each other without the formation of Watson-Crick base pairings between Dk1sub and Dk1enz, thus allowing the DNA phosphorylation reaction. We term this approach proximity-dependent activation of split DNAzyme kinase (ProxSDK). The utility of ProxSDK is demonstrated by engineering a biosensing system that is capable of measuring specific DNA-protein interactions. We envision that the approach described herein will find useful applications in biosensing, imaging, and clinical diagnosis.

From adsorption to crystallization of proteins: Evidence for interface-assisted nucleation.

Protein crystallization is among the key processes in biomolecular research, but the underlying mechanisms are still elusive. Here, we address the role of inevitable interfaces for the nucleation process. Quartz crystal microbalance with dissipation monitoring (QCM-D) with simultaneously optical microscopy, confocal microscopy, and grazing-incidence small angle X-rays scattering (GISAXS) were employed to investigate the temporal behavior from the initial stage of protein adsorption to crystallization. Here we studied the crystallization of the Human Serum Albumin (HSA), the most abundant blood protein, in the presence of a charged surface and a trivalent salt. We found evidence for interface-assisted nucleation of crystals. The kinetic stages involved are initial adsorption followed by enhanced adsorption after longer times, subsequent nucleation, and finally crystal growth. The results highlight the importance of interfaces for protein phase behavior and in particular for nucleation.