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Journal of Agricultural and Food... Sep 2011Soy protein elastomer (SPE) exhibits elastic, extensible, and sticky properties in its native state and displays great potential as an alternative to wheat gluten. The...
Soy protein elastomer (SPE) exhibits elastic, extensible, and sticky properties in its native state and displays great potential as an alternative to wheat gluten. The objective of this study was to better understand the roles of soy protein subunits (polypeptides) contributing to the functional properties of SPE. Six soy protein samples with different subunit compositions were prepared by extracting the proteins at various pH values on the basis of the different solubilities of conglycinin (7S) and glycinin (11S) globulins. Soy protein containing a large amount of high molecular weight aggregates formed from α' and α subunits exhibited stronger viscoelastic solid behavior than other soy protein samples in terms of dynamic elastic and viscous modules. Electrophoresis results revealed that these aggregates are mainly stabilized through disulfide bonds, which also contributed to higher denaturation enthalpy as characterized by DSC and larger size protein aggregates observed by TEM. Besides, the most viscoelastic soy protein sample exhibited flat and smooth surfaces of the protein particles as observed by SEM, whereas other samples had rough and porous particle surfaces. It was proposed that the ability of α' and α to form aggregates and the resultant proper protein-protein interaction in soy proteins are the critical contributions to the continuous network of SPE.
Topics: Chemical Phenomena; Disulfides; Drug Stability; Elasticity; Molecular Weight; Protein Subunits; Solubility; Soybean Proteins; Viscosity
PubMed: 21842907
DOI: 10.1021/jf201077b -
Proteomics Mar 2022Fast, cheap, and easy to implement point-of-care testing for various pathogens constituted a game changer in past years due to its potential for early disease diagnosis....
Fast, cheap, and easy to implement point-of-care testing for various pathogens constituted a game changer in past years due to its potential for early disease diagnosis. Herein, we report on the proof-of-concept of a simple method enabling in vitro detection of a structural spike protein subunit from the SARS-CoV-2 (S ) in aqueous samples. At the core of this discovery lies the well-known paradigm of monitoring the capacitive current across a reconstituted zwitterionic lipid membrane subjected to a periodic transmembrane potential, followed by the real-time spectral analysis enabling the extraction of the second harmonic of the capacitive current. Subsequent changes in the amplitude of this harmonic recorded during lipid membrane-S interactions were correlated with alterations induced in the inner membrane potential profile by the S protein subunit adsorption, and were shown to be augmented by ionic strength, the presence of a specific monoclonal antibody designed against the S subunit and the angiotensin-converting enzyme 2 (ACE2) protein receptor, and uninhibited by the presence of other human serum proteins.
Topics: Biosensing Techniques; COVID-19; Humans; Immunoassay; Lipids; Protein Subunits; SARS-CoV-2; Spike Glycoprotein, Coronavirus
PubMed: 34586750
DOI: 10.1002/pmic.202100047 -
Current Medicinal Chemistry 2008Protein kinase CK2 (formerly referred to as casein kinase II) is an evolutionary conserved, ubiquitous protein kinase. There are two paralog catalytic subunits, i.e.... (Review)
Review
Protein kinase CK2 (formerly referred to as casein kinase II) is an evolutionary conserved, ubiquitous protein kinase. There are two paralog catalytic subunits, i.e. alpha (A1) and alpha' (A2). The alpha and alpha' subunits are linked to two beta subunits to produce a heterotetrameric structure. The catalytic alpha subunits are distantly related to the CMGC subfamily of kinases, such as the Cdk kinases. There are some peculiarities associated with protein kinase CK2, which are not found with most other protein kinases: (i) the enzyme is constitutively active, (ii) it can use ATP and GTP and (iii) it is found elevated in most tumors investigated and rapidly proliferating tissues. With the elucidation of the structure of the catalytic subunit, it was possible to explain why the enzyme is constitutively active [1] and why it can bind GTP [2]. Considerable information on the potential roles of CK2 in various disease processes including cancer has been gained in recent years, and the present review may help to further elucidate its aberrant role in many disease states. Its peculiar structural features [3-9] may be advantageous in designing tailor-made compounds with the possibility to specifically target this protein kinase [10]. Since not all the aspects of what has been published on CK2 can be covered in this review, we would like to recommend the following reviews; (i) for general information on CK2 [11-18] and (ii) with a focus on aberrant CK2 [19-22].
Topics: Casein Kinase II; Humans; Neoplasms; Nervous System Diseases; Protein Subunits; Signal Transduction
PubMed: 18691045
DOI: 10.2174/092986708785132933 -
Progress in Biophysics and Molecular... Oct 2015The V1VO-ATPase (V-ATPase) is the important proton-pump in eukaryotic cells, responsible for pH-homeostasis, pH-sensing and amino acid sensing, and therefore essential... (Review)
Review
The V1VO-ATPase (V-ATPase) is the important proton-pump in eukaryotic cells, responsible for pH-homeostasis, pH-sensing and amino acid sensing, and therefore essential for cell growths and metabolism. ATP-cleavage in the catalytic A3B3-hexamer of V1 has to be communicated via several so-called central and peripheral stalk units to the proton-pumping VO-part, which is membrane-embedded. A unique feature of V1VO-ATPase regulation is its reversible disassembly of the V1 and VO domain. Actin provides a network to hold the V1 in proximity to the VO, enabling effective V1VO-assembly to occur. Besides binding to actin, the 14-subunit V-ATPase interacts with multi-subunit machineries to form cellular sensors, which regulate the pH in cellular compartments or amino acid signaling in lysosomes. Here we describe a variety of subunit-subunit interactions within the V-ATPase enzyme during catalysis and its protein-protein assembling with key cellular machineries, essential for cellular function.
Topics: Adenosine Triphosphatases; Amino Acid Sequence; Animals; Biocatalysis; Eukaryota; Humans; Protein Binding; Protein Subunits; Protons
PubMed: 26033199
DOI: 10.1016/j.pbiomolbio.2015.05.003 -
Microbiology (Reading, England) Oct 2016σ factors are single subunit general transcription factors that reversibly bind core RNA polymerase and mediate gene-specific transcription in bacteria. Previously, an...
σ factors are single subunit general transcription factors that reversibly bind core RNA polymerase and mediate gene-specific transcription in bacteria. Previously, an atypical two-subunit σ factor was identified that activates transcription from a group of related promoters in Bacillus subtilis. Both of the subunits, named SigO and RsoA, share primary sequence similarity with the canonical σ70 family of σ factors and interact with each other and with RNA polymerase subunits. Here we show that the σ70 region 2.3-like segment of RsoA is unexpectedly sufficient for interaction with the amino-terminus of SigO and the β' subunit. A mutational analysis of RsoA identified aromatic residues conserved amongst all RsoA homologues, and often amongst canonical σ factors, that are particularly important for the SigO-RsoA interaction. In a canonical σ factor, region 2.3 amino acids bind non-template strand DNA, trapping the promoter in a single-stranded state required for initiation of transcription. Accordingly, we speculate that RsoA region 2.3 protein-binding activity likely arose from a motif that, at least in its ancestral protein, participated in DNA-binding interactions.
Topics: Amino Acid Sequence; Bacillus subtilis; Bacterial Proteins; Gene Expression Regulation, Bacterial; Promoter Regions, Genetic; Protein Binding; Protein Subunits; Sequence Alignment; Sigma Factor
PubMed: 27558998
DOI: 10.1099/mic.0.000358 -
Cell Structure and Function 2016The Saccharomyces cerevisiae autophagy-initiation complex, Atg1 kinase complex, consists of Atg1, Atg13, Atg17, Atg29, and Atg31, while the corresponding complex in most... (Review)
Review
The Saccharomyces cerevisiae autophagy-initiation complex, Atg1 kinase complex, consists of Atg1, Atg13, Atg17, Atg29, and Atg31, while the corresponding complex in most other eukaryotes, including mammals, is composed of ULK1 (or ULK2), Atg13, FIP200 (also known as RB1CC1), and Atg101. ULKs are homologs of Atg1, and FIP200 is partially homologous to Atg17. However, the sequence of Atg101 is not similar to that of Atg29 or Atg31. Although Atg101 is essential for autophagy and widely conserved in eukaryotes, its precise function and structure have remained largely unknown. Now, structural and cell biological analysis of Atg101 together with its binding partner Atg13 reveal that Atg101 is required for stabilization of "uncapped" Atg13 in most eukaryotes and also for recruitment of downstream Atg proteins through the newly identified WF motif. By contrast, S. cerevisiae has stable "capped" Atg13, which does not require Atg101 for its stabilization. Possible roles for other binding partners such as Atg29, Atg31, and Atg28 in different organisms are also discussed.
Topics: Animals; Autophagy; Humans; Protein Kinases; Protein Subunits; Yeasts
PubMed: 26754330
DOI: 10.1247/csf.15013 -
Chemical Society Reviews Jul 2011Precise knowledge of the three-dimensional structure of a protein is critical, if we are to understand its biological role and mode of action. However, today it is... (Review)
Review
Precise knowledge of the three-dimensional structure of a protein is critical, if we are to understand its biological role and mode of action. However, today it is becoming increasingly clear that dissecting the protein's structural architecture is not enough: a complete description of biomolecular activity must also include the dimension of time. Protein motion and dynamics are crucial for protein stability and reactivity. A range of techniques have been developed for probing dynamic processes. In this tutorial review, we focus on one of these approaches--structural mass spectrometry (MS). MS has the ability to capture functional conformational transitions in the slow time regime, from a few milliseconds to hours. The power of this approach lies not only in its sensitivity and speed of analysis, but also in the fact that it is a non-ensemble technique. Thus, within a single spectrum, the entire distribution of co-existing states can be resolved. In discussing the challenges, advantages and limitations of the field, as well as future directions, we highlight the applicability of MS for quantitative monitoring of structural kinetics. In particular, we describe the array of MS-based strategies that are available for capturing protein folding, enzymatic reactions, ligand interactions, subunit exchange and biogenesis pathways.
Topics: Kinetics; Mass Spectrometry; Protein Conformation; Protein Folding; Protein Subunits; Proteins
PubMed: 21547331
DOI: 10.1039/c1cs15052a -
Chemphyschem : a European Journal of... Mar 2014The limit of subdiffraction imaging with fluorescent proteins currently lies at 20 nm, and therefore most protein complexes are too small (2-5 nm) to spatially resolve... (Review)
Review
The limit of subdiffraction imaging with fluorescent proteins currently lies at 20 nm, and therefore most protein complexes are too small (2-5 nm) to spatially resolve their individual subunits by optical means. However, the number and stoichiometry of subunits within an immobilized protein complex can be resolved by the observation of photobleaching steps of individual fluorophores or co-localization of single-molecule fluorescence emission in multiple colors. We give an overview of the proteins that have been investigated by this approach and the different techniques that can be used to immobilize and label the proteins. This minireview should serve as a guideline for scientists who want to employ single-molecule subunit counting for their research.
Topics: Fluorescence; Luminescent Proteins; Photobleaching; Protein Subunits
PubMed: 24481650
DOI: 10.1002/cphc.201301092 -
Cellular and Molecular Life Sciences :... Dec 2014In eukaryotic cells, proteasomes are highly conserved protease complexes and eliminate unwanted proteins which are marked by poly-ubiquitin chains for degradation. The... (Review)
Review
In eukaryotic cells, proteasomes are highly conserved protease complexes and eliminate unwanted proteins which are marked by poly-ubiquitin chains for degradation. The 26S proteasome consists of the proteolytic core particle, the 20S proteasome, and the 19S regulatory particle, which are composed of 14 and 19 different subunits, respectively. Proteasomes are the second-most abundant protein complexes and are continuously assembled from inactive precursor complexes in proliferating cells. The modular concept of proteasome assembly was recognized in prokaryotic ancestors and applies to eukaryotic successors. The efficiency and fidelity of eukaryotic proteasome assembly is achieved by several proteasome-dedicated chaperones that initiate subunit incorporation and control the quality of proteasome assemblies by transiently interacting with proteasome precursors. It is important to understand the mechanism of proteasome assembly as the proteasome has key functions in the turnover of short-lived proteins regulating diverse biological processes.
Topics: Animals; Humans; Models, Biological; Models, Molecular; Proteasome Endopeptidase Complex; Protein Binding; Protein Conformation; Protein Subunits
PubMed: 25107634
DOI: 10.1007/s00018-014-1699-8 -
Journal of Chromatography. A Apr 2014Tag-free proteins or protein complexes represent certainly the most authentic starting points for functional or structural studies. They can be obtained by conventional...
Tag-free proteins or protein complexes represent certainly the most authentic starting points for functional or structural studies. They can be obtained by conventional multi-step chromatography from native or recombinant tag-free sources. Alternatively, they can be expressed and purified using a cleavable N-terminal affinity tag that is subsequently removed by a site-specific protease. Proteolytic tag-removal can also be performed "on-column". We show here that this not only represents a very efficient workflow, but also drastically improves the purity of the resulting protein preparations. Precondition for effective on-column-cleavage is, however, that the tag-cleaving protease does not bind the stationary phase. We introduce scAtg4 and xlUsp2 as very good and bdSENP1, bdNEDP1 as well as ssNEDP1 as ideal proteases for on-column cleavage at 4°C. Four of these proteases (bdSENP1, bdNEDP1, scAtg4, xlUsp2) as well as TEV protease display orthogonal, i.e. mutually exclusive cleavage specificities. We combined these features into a streamlined method for the production of highly pure protein complexes: Orthogonal affinity tags and protease recognitions modules are fused to individual subunits. Following co-expression or in-vitro complex assembly, consecutive cycles of affinity capture and proteolytic release then select sequentially for the presence of each orthogonally tagged subunit, yielding protein complexes of well-defined subunit stoichiometry.
Topics: Chromatography, Affinity; Histidine; Multiprotein Complexes; Peptide Hydrolases; Protein Subunits; Recombinant Proteins; SUMO-1 Protein; Substrate Specificity
PubMed: 24636567
DOI: 10.1016/j.chroma.2014.02.030