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Specificity of interactions between two DNA strands, or between protein and DNA, is often achieved by varying bases or side chains coming off the DNA or protein backbone—for example, the bases participating in Watson–Crick pairing in the double helix, or the side chains contacting DNA in TALEN–DNA complexes. By contrast, specificity of protein–protein interactions usually involves backbone shape complementarity 1 , which is less modular and hence harder to generalize. Coiled-coil heterodimers are an exception, but the restricted geometry of interactions across the heterodimer interface (primarily at the heptad a and d positions 2 ) limits the number of orthogonal pairs that can be created simply by varying side-chain interactions 3 , 4 . Here we show that protein–protein interaction specificity can be achieved using extensive and modular side-chain hydrogen-bond networks. We used the Crick generating equations 5 to produce millions of four-helix backbones with varying degrees of supercoiling around a central axis, identified those accommodating extensive hydrogen-bond networks, and used Rosetta to connect pairs of helices with short loops and to optimize the remainder of the sequence. Of 97 such designs expressed in Escherichia coli , 65 formed constitutive heterodimers, and the crystal structures of four designs were in close agreement with the computational models and confirmed the designed hydrogen-bond networks. In cells, six heterodimers were fully orthogonal, and in vitro—following mixing of 32 chains from 16 heterodimer designs, denaturation in 5 M guanidine hydrochloride and reannealing—almost all of the interactions observed by native mass spectrometry were between the designed cognate pairs. The ability to design orthogonal protein heterodimers should enable sophisticated protein-based control logic for synthetic biology, and illustrates that nature has not fully explored the possibilities for programmable biomolecular interaction modalities. Computational design incorporating modular buried hydrogen networks produces highly orthogonal protein heterodimers.

Dry molten globular (DMG) intermediates, an expanded form of the native protein with a dry core, have been observed during denaturant-induced unfolding of many proteins. These observations are counterintuitive because traditional models of chemical denaturation rely on changes in solvent-accessible surface area, and there is no notable change in solvent-accessible surface area during the formation of the DMG. Here we show, using multisite fluorescence resonance energy transfer, far-UV CD, and kinetic thiol-labeling experiments, that the guanidinium chloride (GdmCl)-induced unfolding of RNase H also begins with the formation of the DMG. Population of the DMG occurs within the 5-ms dead time of our measurements. We observe that the size and/or population of the DMG is linearly dependent on [GdmCl], although not as strongly as the second and major step of unfolding, which is accompanied by core solvation and global unfolding. This rapid GdmCl-dependent population of the DMG indicates that GdmCl can interact with the protein before disrupting the hydrophobic core. These results imply that the effect of chemical denaturants cannot be interpreted solely as a disruption of the hydrophobic effect and strongly support recent computational studies, which hypothesize that chemical denaturants first interact directly with the protein surface before completely unfolding the protein in the second step (direct interaction mechanism).

Herein, we discuss changes in the emulsifying properties of myofibrillar protein (MP) because of protein denaturation and aggregation from quick-frozen pork patties with multiple fat levels and freeze–thaw (F–T) cycles. Protein denaturation and aggregation were confirmed by the significantly increased surface hydrophobicity, turbidity, and particle size, as well as the significantly decreased solubility and absolute zeta potential, of MPs with increases in fat levels and F–T cycles (p < 0.05). After multiple F–T cycles, the emulsifying activity and emulsion stability indices of all samples were significantly reduced (p < 0.05). The emulsion droplets of MP increased in size, and their distributions were dense and irregular. The results demonstrated that protein denaturation and aggregation due to multiple F–T cycles and fat levels changed the distribution of surface chemical groups and particle sizes of protein, thus affecting the emulsifying properties.

Both gamma rays and atmospheric pressure plasma are known to have anticancer properties. While their mechanism actions are still not clear, in some contexts they work in similar manner, while in other contexts they work differently. So to understand these relationships, we have studied Myoglobin protein after the treatment of gamma rays and dielectric barrier discharge (DBD) plasma, and analyzed the changes in thermodynamic properties and changes in the secondary structure of protein after both treatments. The thermodynamic properties were analyzed using chemical and thermal denaturation after both treatments. We have also studied the action of gamma rays and DBD plasma on myoglobin in the presence of osmolytes, such as sorbitol and trehalose. For deep understanding of the action of gamma rays and DBD plasma, we have analyzed the reactive species generated by them in buffer at all treatment conditions. Finally, we have used molecular dynamic simulation to understand the hydrogen peroxide action on myoglobin with or without osmolytes, to gain deeper insight into how the osmolytes can protect the protein structure from the reactive species generated by gamma rays and DBD plasma.

The article proposes using Computational Fluid Dynamics to predict protein denaturation and weight loss in pork loin samples during baking. The optimal conditions of heat treatment were determined to be at 181.55℃, 20% humidity, and a fan speed of 639.18 rpm. Following this process, the denaturation of various pork loin proteins was measured, and the weight loss was determined to be 12.21 ± 0.89%. In the laboratory study at the verification stage, it was found that myosin and actin denaturation and body weight loss did not differ significantly from the values predicted on the basis of the response surface model developed using simulation data. However, the degree of sarcoplasmic protein denaturation observed in the laboratory was lower than the model's predictions, while collagen denaturation was higher. The study suggests that computational fluid dynamics can be a useful tool for predicting protein denaturation and weight loss in pork loin samples and could improve product quality and consistency in the food industry.

Ultrasound (US) application on milk fat homogenization was compared to conventional treatment by pressure in terms of milk fat globule (MFG) size, fermentation process kinetics, and viscosity of set type yogurt. Homogenization of milk by (i) US (frequency 20 kHz, amplitude 150–750 W) and (ii) two-stage pressure (10–30 MPa/5 MPa) was examined.The more intense the homogenization was, the smaller the MFG became, regardless of the applied method; high-intensity US homogenization reduced the MFG size to 0.78 μm. The fermentation kinetics of ultrasonicated milk samples were significantly different to the samples homogenized by pressure in terms of pH and viscosity. The pH reduction rate and the duration of pH lag phase of US homogenized milk were significantly lower (42 % for μₚHand 52 % for λₚH) compared to those of milk homogenized by pressure. In terms of viscosity evolution, the US homogenization leads to increased rates of increase (by up to 64 %) and shorter lag phases (by up to 56 %), compared to pressure homogenization. Yogurt coagulum obtained at the end of the fermentation (pH = 4.6) of milk homogenized by US had significantly higher viscosity values compared to those of milk homogenized with pressure. The difference in the evolution and the end values of yogurt’s viscosity was attributed to the denaturation of milk proteins occurring during the US treatment of the milk. US treatment of milk leads to decrease of soluble protein content and composition, which is possibly connected with the formation of insoluble high molecular weight coaggregates that occurred due to whey protein denaturation during US treatment.

A useful tool to analyze the ligands and/or environmental contribution to protein stability is represented by the Synchrotron Radiation Circular Dichroism UV-denaturation assay that consists in the acquisition of several consecutive repeated far-UV SRCD spectra. Recently we demonstrated that the prevailing mechanism of this denaturation involves the generation of free radicals and reactive oxygen species (ROS). In this work, we analyzed the effect of buffering agents commonly used in spectroscopic measurements, including MOPS (3-(N-morpholino) propanesulfonic acid), HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid), TRIS-HCl (tris-hydroxymethil aminomethane hydrochloride), and phosphate, on the efficiency of protein denaturation caused by exposure to UV radiation. Fluorescence experiments confirmed the presence of ROS and were used to determine the rate of ROS generation. Our results indicate that the efficiency of the denaturation process is strongly influenced by the buffer composition with MOPS and HEPES acting also as scavengers and that the presence of proteins itself influenced the ROS formation rate.

It was shown that at low concentrations guanidine hydrochloride (GdnHCl) can cause aggregation of proteins in partially folded state and that fluorescent dye 1-anilinonaphthalene-8-sulfonic acid (ANS) binds with these aggregates rather than with hydrophobic clusters on the surface of protein in molten globule state. That is why the increase in ANS fluorescence intensity is often recorded in the pathway of protein denaturation by GdnHCl, but not by urea. So what was previously believed to be the molten globule state in the pathway of protein denaturation by GdnHCl, in reality, for some proteins represents the aggregates of partially folded molecules.

Arid ecosystems constitute a promising source of actinobacteria producing new bioactive molecules. This study aimed to explore different biological activities of actinomycetes isolated from the rhizosphere of Phoenix dactylifera L. in the Ghardaia region, Algeria. A total of 18 actinobacteria were isolated and studied for their enzymatic and antimicrobial activities. All isolates shared cellulase and catalase activity; most of them produced amylase (94%), esterase (84%), lecithinase and lipoproteins (78%), caseinase (94%), and gelatinase (72%). The isolates could coagulate (56%) or peptonize (28%) skim milk. Overall, 72% of the isolates exhibited significant antibacterial activity against at least one test bacteria, while 56% demonstrated antifungal activity against at least one test fungi. Based on enzyme production and antimicrobial activity, isolate SGI16 was selected for secondary metabolite extraction by ethyl acetate. The crude extract of SGI16 was analyzed using DPPH and BSA denaturation inhibition tests, revealing significant antioxidant power (IC50 = 7.24 ± 0.21 μg mL−1) and protein denaturation inhibitory capacity (IC50 = 492.41 ± 0.47 μg mL−1). Molecular identification based on 16S rDNA analysis showed that SGI16 belonged to the genus Streptomyces. The findings highlight that date palms’ rhizosphere actinobacteria are a valuable source of biomolecules of biotechnological interest.

In this study, the effect of various heating temperatures (61–70 °C) and times (1–10 min) on physical and chemical properties of liquid egg yolk (LEY) and mayonnaise were investigated. Initially, we found that the increase of LEY protein denaturation was highly correlated with the increase of temperature and time, without causing either protein degradation or aggregation. In addition, the viscosity and particle size of LEY were significantly increased with greater heating temperature and time. Furthermore, the emulsification stability of mayonnaise prepared from thermally processed LEY were significantly better than that of the unheated control group, in particular, the emulsion stability of mayonnaise was higher at a temperature ranging from 62 °C to 68 °C, whereas the emulsion stability decreased above 69 °C. A rheological analysis showed that mayonnaise prepared from thermally processed LEY has higher shear stress when compared with the control group. Indeed, a sharp increase in the shear stress was observed when LEY was heated above 67 °C. Results from storage behavior analysis suggest that mayonnaise prepared from thermally processed LEY failed to affect the chemical qualities of mayonnaise, as evidenced by the fact that acid values and TBA values were not statistically significant with the unheated control group. Microscopic observation indicates that the number of complete oil droplets were significantly reduced at higher heating (70 °C/5 and 10 min) conditions. Finally, the sensory evaluation results suggest that mayonnaise prepared from thermally processed LEY does not influence the appearance, aroma, taste, greasy feeling, and overall acceptance of mayonnaise, as indicated by there being no significant differences between the experimental group and the control group (p > 0.05). We conclude from our study that a combination of heating conditions over 67 °C/5 min can allow the mayonnaise to retain better quality in terms of stability.