Membrane proteins are a large, diverse group of proteins, serving a multitude of cellular functions. They are difficult to study because of their requirement of a lipid membrane for function. Here we show that two-photon polarization microscopy can take advantage of the cell membrane requirement to yield insights into membrane protein structure and function, in living cells and organisms. The technique allows sensitive imaging of G-protein activation, changes in intracellular calcium concentration and other processes, and is not limited to membrane proteins. Conveniently, many suitable probes for two-photon polarization microscopy already exist.

• MeSH
• buněčná membrána metabolismus   ultrastruktura   MeSH
• konformace proteinů MeSH
• membránové proteiny metabolismus   ultrastruktura   MeSH
• mikroskopie fluorescenční multifotonová metody   MeSH
• polarizační mikroskopie metody   MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• membránové proteiny MeSH

Chemical cross-linking coupled with mass spectrometry is a popular technique for deriving structural information on proteins and protein complexes. Also, cross-linking has become a powerful tool for stabilizing macromolecular complexes for single-particle cryo-electron microscopy. However, an effect of cross-linking on protein structure and function should not be forgotten, and surprisingly, it has not been investigated in detail so far. Here, we used kinetic studies, mass spectrometry, and NMR spectroscopy to systematically investigate an impact of cross-linking on structure and function of human carbonic anhydrase and alcohol dehydrogenase 1 from Saccharomyces cerevisiae. We found that cross-linking induces rather local structural disturbances and the overall fold is preserved even at a higher cross-linker concentration. The results establish general experimental conditions for chemical cross-linking with minimal effect on protein structure and function.

• MeSH
• alkoholdehydrogenasa chemie   MeSH
• hmotnostní spektrometrie MeSH
• karboanhydrasy chemie   MeSH
• konformace proteinů MeSH
• lidé MeSH
• molekulární modely MeSH
• multimerizace proteinu MeSH
• nukleární magnetická rezonance biomolekulární MeSH
• reagencia zkříženě vázaná chemie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• alkoholdehydrogenasa MeSH
• karboanhydrasy MeSH
• reagencia zkříženě vázaná MeSH

Quantitative structure-function relationships (QSFR) and quantitative structure-stability relationships (QSSR) analyses are described here. The objective of these analyses is to investigate and quantitatively describe the effect of the changes in structure of protein on its function or stability. During the analysis, the structural and physico-chemical properties of the amino acid residues are related to activity or stability data derived for the group of proteins containing systematic substitutions at certain positions. Four examples of the application of these analyses on the data obtained with proteins modified by site-directed mutagenesis experiments are provided. Structure-function relationships were studied for 15 mutants in position 172 of the haloalkane dehalogenase and 19 mutants in position 222 of the subtilisin, while the structure-stability relationships were investigated for 13 mutants in position 157 of phage T4 lysozyme and 18 mutants in position 49 of alpha-subunits tryptophan synthase. A total of 402 molecular descriptors derived from AAindex database were used to quantify amino acid properties and the multivariate statistical technique--partial least squares projections to latent structures--was used to identify those of them which are important for explanation of the activity and stability data. Quantitative models were developed and internally validated for every data set. The possibilities for further development of both analyses and their application for predictive and analytical purposes in protein engineering research are discussed.

• MeSH
• kyselina glutamová chemie   MeSH
• mutageneze cílená MeSH
• proteinové inženýrství MeSH
• termodynamika MeSH
• tryptofansynthasa chemie   genetika   MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• kyselina glutamová MeSH
• tryptofansynthasa MeSH

Comparative evolutionary genomics has revealed that novel protein coding genes can emerge randomly from non-coding DNA. While most of the myriad of transcripts which continuously emerge vanish rapidly, some attain regulatory regions, become translated and survive. More surprisingly, sequence properties of de novo proteins are almost indistinguishable from randomly obtained sequences, yet de novo proteins may gain functions and integrate into eukaryotic cellular networks quite easily. We here discuss current knowledge on de novo proteins, their structures, functions and evolution. Since the existence of de novo proteins seems at odds with decade-long attempts to construct proteins with novel structures and functions from scratch, we suggest that a better understanding of de novo protein evolution may fuel new strategies for protein design.

• MeSH
• genomika MeSH
• molekulární evoluce * MeSH
• proteiny * genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• proteiny * MeSH

The ability to predict and design protein structures has led to numerous applications in medicine, diagnostics and sustainable chemical manufacture. In addition, the wealth of predicted protein structures has advanced our understanding of how life's molecules function and interact. Honouring the work that has fundamentally changed the way scientists research and engineer proteins, the Nobel Prize in Chemistry in 2024 was awarded to David Baker for computational protein design and jointly to Demis Hassabis and John Jumper, who developed AlphaFold for machine-learning-based protein structure prediction. Here, we highlight notable contributions to the development of these computational tools and their importance for the design of functional proteins that are applied in organic synthesis. Notably, both technologies have the potential to impact drug discovery as any therapeutic protein target can now be modelled, allowing the de novo design of peptide binders and the identification of small molecule ligands through in silico docking of large compound libraries. Looking ahead, we highlight future research directions in protein engineering, medicinal chemistry and material design that are enabled by this transformative shift in protein science.

• Klíčová slova
• AlphaFold, Computational protein design, Nobel prize, Protein engineering, Protein structure prediction,
• MeSH
• biokatalýza MeSH
• konformace proteinů MeSH
• proteinové inženýrství MeSH
• proteiny * chemie   metabolismus   MeSH
• strojové učení MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• proteiny * MeSH

In a woman suffering from IgE myeloma, hay fever and polyvalent respiratory and skin allergy the IgE monoclonal protein VL was isolated and investigated with respect to structural and functional properties. The amino acid sequence of 22 isolated peptides--especially of the biologically significant C2-C3 part--corresponded with that originally described by Bennich et al. (Immunol Rev 1978;41:3-23; Prog Immunol 1974;13:49-58). However, in mass spectrometry the sugar residues on ASN 99 (219) and 252 (371) were deficient in sialic acids. The native IgE VL protein precipitated with high intensity all mannose-specific lectins as concanavalin A (Con A) and was able to release histamine after triggering by these lectins. The same lectins also elicited more histamine release and more positive skin reactions in atopic than in healthy persons. In sera from atopic patients the binding of IgE on Con A Sepharose 4B column was stronger than in normal persons. It is suggested that changes in the IgE glycosylation state may contribute to IgE-mediated pictures of clinical allergy by the nonimmunological pathway.

• MeSH
• alergeny chemie   MeSH
• časná přecitlivělost imunologie   MeSH
• glykosylace MeSH
• imunoglobulin E krev   chemie   fyziologie   MeSH
• imunoglobuliny - kappa-řetězce chemie   izolace a purifikace   fyziologie   MeSH
• lektiny chemie   MeSH
• lidé MeSH
• mnohočetný myelom imunologie   MeSH
• molekulární sekvence - údaje MeSH
• molekulová hmotnost MeSH
• myelomové proteiny chemie   izolace a purifikace   MeSH
• oligosacharidy chemie   MeSH
• precipitinové testy MeSH
• sefarosa analogy a deriváty   chemie   MeSH
• sekvence aminokyselin MeSH
• variabilní oblast imunoglobulinu chemie   izolace a purifikace   fyziologie   MeSH
• vazebná místa protilátek MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Check Tag
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• alergeny MeSH
• concanavalin A-sepharose MeSH Prohlížeč
• imunoglobulin E MeSH
• imunoglobuliny - kappa-řetězce MeSH
• lektiny MeSH
• myelomové proteiny MeSH
• oligosacharidy MeSH
• sefarosa MeSH
• variabilní oblast imunoglobulinu MeSH

The actin family members, consisting of actin and actin-related proteins (ARPs), are essential components of chromatin remodeling complexes. ARP6, one of the nuclear ARPs, is part of the Snf-2-related CREB-binding protein activator protein (SRCAP) chromatin remodeling complex, which promotes the deposition of the histone variant H2A.Z into the chromatin. In this study, we showed that ARP6 influences the structure and the function of the nucleolus. ARP6 is localized in the central region of the nucleolus, and its knockdown induced a morphological change in the nucleolus. We also found that in the presence of high concentrations of glucose ARP6 contributed to the maintenance of active ribosomal DNA (rDNA) transcription by placing H2A.Z into the chromatin. In contrast, under starvation, ARP6 was required for cell survival through the repression of rDNA transcription independently of H2A.Z. These findings reveal novel pleiotropic roles for the actin family in nuclear organization and metabolic homeostasis.

• Klíčová slova
• ARP6, Actin-related protein, Histone H2A.Z, Nucleolus, Wndchrm,
• MeSH
• adenosintrifosfatasy metabolismus   MeSH
• aktiny metabolismus   fyziologie   MeSH
• buněčné jadérko metabolismus   fyziologie   MeSH
• chromozomální proteiny, nehistonové metabolismus   fyziologie   MeSH
• genetická transkripce fyziologie   MeSH
• glukosa metabolismus   MeSH
• HeLa buňky MeSH
• lidé MeSH
• ribozomální DNA genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Intramural MeSH
• Názvy látek
• ACTR6 protein, human MeSH Prohlížeč
• adenosintrifosfatasy MeSH
• aktiny MeSH
• chromozomální proteiny, nehistonové MeSH
• glukosa MeSH
• ribozomální DNA MeSH
• SRCAP protein, human MeSH Prohlížeč

Many aspects of protein function regulation require specific protein-protein interactions to carry out the exact biochemical and cellular functions. The highly conserved members of the 14-3-3 protein family mediate such interactions and through binding to hundreds of other proteins provide multitude of regulatory functions, thus playing key roles in many cellular processes. The 14-3-3 protein binding can affect the function of the target protein in many ways including the modulation of its enzyme activity, its subcellular localization, its structure and stability, or its molecular interactions. In this minireview, we focus on mechanisms of the 14-3-3 protein-dependent regulation of three important 14-3-3 binding partners: yeast neutral trehalase Nth1, regulator of G-protein signaling 3 (RGS3), and phosducin.

• MeSH
• DNA-glykosylasy chemie   ultrastruktura   MeSH
• DNA-lyasa (apurinová nebo apyrimidinová) chemie   ultrastruktura   MeSH
• fosfoproteiny chemie   ultrastruktura   MeSH
• konformace proteinů MeSH
• lidé MeSH
• molekulární sekvence - údaje MeSH
• multienzymové komplexy chemie   ultrastruktura   MeSH
• oční proteiny chemie   ultrastruktura   MeSH
• proteiny 14-3-3 chemie   ultrastruktura   MeSH
• proteiny RGS chemie   ultrastruktura   MeSH
• proteiny vázající GTP - regulátory chemie   ultrastruktura   MeSH
• Schizosaccharomyces pombe - proteiny chemie   ultrastruktura   MeSH
• sekvence aminokyselin MeSH
• vazba proteinů MeSH
• vazebná místa MeSH
• vztahy mezi strukturou a aktivitou MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• DNA-glykosylasy MeSH
• DNA-lyasa (apurinová nebo apyrimidinová) MeSH
• fosfoproteiny MeSH
• multienzymové komplexy MeSH
• Nth1 protein, S pombe MeSH Prohlížeč
• oční proteiny MeSH
• phosducin MeSH Prohlížeč
• proteiny 14-3-3 MeSH
• proteiny RGS MeSH
• proteiny vázající GTP - regulátory MeSH
• RGS3 protein, human MeSH Prohlížeč
• Schizosaccharomyces pombe - proteiny MeSH

BACKGROUND: Protein function is determined by many factors, namely by its constitution, spatial arrangement, and dynamic behavior. Studying these factors helps the biochemists and biologists to better understand the protein behavior and to design proteins with modified properties. One of the most common approaches to these studies is to compare the protein structure with other molecules and to reveal similarities and differences in their polypeptide chains. RESULTS: We support the comparison process by proposing a new visualization technique that bridges the gap between traditionally used 1D and 3D representations. By introducing the information about mutual positions of protein chains into the 1D sequential representation the users are able to observe the spatial differences between the proteins without any occlusion commonly present in 3D view. Our representation is designed to serve namely for comparison of multiple proteins or a set of time steps of molecular dynamics simulation. CONCLUSIONS: The novel representation is demonstrated on two usage scenarios. The first scenario aims to compare a set of proteins from the family of cytochromes P450 where the position of the secondary structures has a significant impact on the substrate channeling. The second scenario focuses on the protein flexibility when by comparing a set of time steps our representation helps to reveal the most dynamically changing parts of the protein chain.

• Klíčová slova
• Molecular sequence analysis, Molecular structure and function, Molecular visualization,
• MeSH
• algoritmy MeSH
• molekulární modely MeSH
• proteiny chemie   MeSH
• sekundární struktura proteinů * MeSH
• sekvence aminokyselin MeSH
• sekvenční seřazení MeSH
• simulace molekulární dynamiky * MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• proteiny MeSH

Regulator of G protein signaling (RGS) proteins function as GTPase-activating proteins for the α-subunit of heterotrimeric G proteins. The function of certain RGS proteins is negatively regulated by 14-3-3 proteins, a family of highly conserved regulatory molecules expressed in all eukaryotes. In this study, we provide a structural mechanism for 14-3-3-dependent inhibition of RGS3-Gα interaction. We have used small angle x-ray scattering, hydrogen/deuterium exchange kinetics, and Förster resonance energy transfer measurements to determine the low-resolution solution structure of the 14-3-3ζ·RGS3 complex. The structure shows the RGS domain of RGS3 bound to the 14-3-3ζ dimer in an as-yet-unrecognized manner interacting with less conserved regions on the outer surface of the 14-3-3 dimer outside its central channel. Our results suggest that the 14-3-3 protein binding affects the structure of the Gα interaction portion of RGS3 as well as sterically blocks the interaction between the RGS domain and the Gα subunit of heterotrimeric G proteins.

• MeSH
• cirkulární dichroismus MeSH
• fosforylace MeSH
• hmotnostní spektrometrie MeSH
• lidé MeSH
• maloúhlový rozptyl MeSH
• proteiny 14-3-3 chemie   genetika   metabolismus   MeSH
• proteiny aktivující GTPasu chemie   genetika   metabolismus   MeSH
• proteiny RGS MeSH
• proteiny vázající GTP chemie   genetika   metabolismus   MeSH
• rezonanční přenos fluorescenční energie MeSH
• sekundární struktura proteinů MeSH
• signální transdukce MeSH
• terciární struktura proteinů MeSH
• vazba proteinů MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• proteiny 14-3-3 MeSH
• proteiny aktivující GTPasu MeSH
• proteiny RGS MeSH
• proteiny vázající GTP MeSH
• RGS3 protein, human MeSH Prohlížeč