Preparation for Optimal Conformation of Lysozyme with Nanodiamond and Nanosilica as Carriers

2010 ◽  
Vol 28 (12) ◽  
pp. 2520-2526 ◽  
Author(s):  
Victor Wei-Keh Wu ◽  
Fumi KureKo
Keyword(s):  
Optimal Conformation

scholarly journals Assembly of the Drosophila 26 S proteasome is accompanied by extensive subunit rearrangements

2002 ◽  
Vol 365 (2) ◽  
pp. 527-536 ◽  
Author(s):  
Éva KURUCZ ◽  
István ANDÓ ◽  
Máté SÜMEGI ◽  
Harald HÖLZL ◽  
Barbara KAPELARI ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
26S Proteasome ◽  
Cross Linking ◽  
20S Proteasome ◽  
Electron Microscopic ◽  
Immunoblot Assay ◽  
Optimal Conformation ◽  
Regulatory Complex ◽  
Atpase Subunits ◽  
The Cross

The subunit contacts in the regulatory complex of the Drosophila 26 S proteasome were studied through the cross-linking of closely spaced subunits of the complex, and analysis of the cross-linking pattern in an immunoblot assay with the use of subunit-specific monoclonal antibodies. The cross-linking pattern of the purified 26 S proteasome exhibits significant differences as compared with that of the purified free regulatory complex. It is shown that the observed differences are due to extensive rearrangement of the subunit contacts accompanying the assembly of the 26 S proteasome from the regulatory complex and the 20S proteasome. Cross-linking studies and electron microscopic examinations revealed that these changes are reversible and follow the assembly or the disassembly of the 26 S proteasome. Although the majority of the changes observed in the subunit contacts affected the hexameric ring of the ATPase subunits, the alterations extended over the whole of the regulatory complex, affecting subunit contacts even in the lid subcomplex. Changes in the subunit contacts, similar to those in the regulatory complex, were detected in the 20S proteasome. These observations indicate that the assembly of the 26 S proteasome is not simply a passive docking of two rigid subcomplexes. In the course of the assembly, the interacting subcomplexes mutually rearrange their structures so as to create the optimal conformation required for the assembly and the proper functioning of the 26S proteasome.

scholarly journals Structural insights into the promutagenic bypass of the major cisplatin-induced DNA lesion

2020 ◽  
Vol 477 (5) ◽  
pp. 937-951
Author(s):  
Hala Ouzon-Shubeita ◽  
Caroline K. Vilas ◽  
Seongmin Lee
Keyword(s):  
Dna Polymerase ◽  
Active Site ◽  
Cisplatin Resistance ◽  
Structural Basis ◽  
Abasic Site ◽  
Dna Polymerase Β ◽  
Dna Lesion ◽  
Human Dna ◽  
Optimal Conformation ◽  
Structural Insights

The cisplatin-1,2-d(GpG) (Pt-GG) intrastrand cross-link is the predominant DNA lesion generated by cisplatin. Cisplatin has been shown to predominantly induce G to T mutations and Pt-GG permits significant misincorporation of dATP by human DNA polymerase β (polβ). In agreement, polβ overexpression, which is frequently observed in cancer cells, is linked to cisplatin resistance and a mutator phenotype. However, the structural basis for the misincorporation of dATP opposite Pt-GG is unknown. Here, we report the first structures of a DNA polymerase inaccurately bypassing Pt-GG. We solved two structures of polβ misincorporating dATP opposite the 5′-dG of Pt-GG in the presence of Mg2+ or Mn2+. The Mg2+-bound structure exhibits a sub-optimal conformation for catalysis, while the Mn2+-bound structure is in a catalytically more favorable semi-closed conformation. In both structures, dATP does not form a coplanar base pairing with Pt-GG. In the polβ active site, the syn-dATP opposite Pt-GG appears to be stabilized by protein templating and pi stacking interactions, which resembles the polβ-mediated dATP incorporation opposite an abasic site. Overall, our results suggest that the templating Pt-GG in the polβ active site behaves like an abasic site, promoting the insertion of dATP in a non-instructional manner.

scholarly journals Sequential activation of human signal recognition particle by the ribosome and signal sequence drives efficient protein targeting

2018 ◽  
Vol 115 (24) ◽  
pp. E5487-E5496 ◽  
Author(s):  
Jae Ho Lee ◽  
Sowmya Chandrasekar ◽  
SangYoon Chung ◽  
Yu-Hsien Hwang Fu ◽  
Demi Liu ◽  
...  
Keyword(s):  
Single Molecule ◽  
Targeted Delivery ◽  
Protein Targeting ◽  
Signal Sequence ◽  
Signal Recognition Particle ◽  
Signal Recognition ◽  
Optimal Conformation ◽  
Sequential Mechanism ◽  
Sequential Activation ◽  
Sense And Respond

Signal recognition particle (SRP) is a universally conserved targeting machine that mediates the targeted delivery of ∼30% of the proteome. The molecular mechanism by which eukaryotic SRP achieves efficient and selective protein targeting remains elusive. Here, we describe quantitative analyses of completely reconstituted human SRP (hSRP) and SRP receptor (SR). Enzymatic and fluorescence analyses showed that the ribosome, together with a functional signal sequence on the nascent polypeptide, are required to activate SRP for rapid recruitment of the SR, thereby delivering translating ribosomes to the endoplasmic reticulum. Single-molecule fluorescence spectroscopy combined with cross-complementation analyses reveal a sequential mechanism of activation whereby the ribosome unlocks the hSRP from an autoinhibited state and primes SRP to sample a variety of conformations. The signal sequence further preorganizes the mammalian SRP into the optimal conformation for efficient recruitment of the SR. Finally, the use of a signal sequence to activate SRP for receptor recruitment is a universally conserved feature to enable efficient and selective protein targeting, and the eukaryote-specific components confer upon the mammalian SRP the ability to sense and respond to ribosomes.

scholarly journals Nebulette is a powerful cytolinker organizing desmin and actin in mouse hearts

2016 ◽  
Vol 27 (24) ◽  
pp. 3869-3882 ◽  
Author(s):  
Daniel A. Hernandez ◽  
Christina M. Bennett ◽  
Lyubov Dunina-Barkovskaya ◽  
Tatjana Wedig ◽  
Yassemi Capetanaki ◽  
...  
Keyword(s):  
Functional Relationship ◽  
Striated Muscle ◽  
Muscle Relaxation ◽  
Canine Heart ◽  
Filament Assembly ◽  
Optimal Conformation ◽  
Assembly Kinetics ◽  
In Vitro Interaction ◽  
Fine Tune

In the hearts of patients bearing nebulette mutations, a severe general disorganization in cardiomyocytes of the extrasarcomeric desmin intermediate filament system is frequently observed. However, the molecular and functional relationship between the desmin cytoskeleton and nebulette-containing sarcomeres is still unclear. Here we report a high-affinity in vitro interaction between nebulette and desmin filaments. A major interaction site has been mapped to the desmin α-helical rod domain, indicating that the filament core is directly involved in the binding of nebulette. The disease-mutant desmin variants E245D and T453I exhibited increased binding affinity for nebulette, delayed filament assembly kinetics, and caused significant weakening of networks. In isolated chick cardiomyocytes and sections from canine heart, we revealed by ground-state depletion and confocal microscopies that module 5 of nebulette extends outward from Z-disk–associated desmin filaments toward the center of the sarcomere. Accordingly, in the myocardium of Des−/− mice, elevated levels of cardiac actin correlated with alterations in the distribution of nebulette. Our data suggest that a well-organized desmin network is required to accommodate an optimal conformation of nebulette on sarcomeres to bind and recruit cardiac α-actin. Hence we propose that nebulette acts in synergy with nebulin to reinforce and temporally fine-tune striated muscle relaxation–contraction cycles.

scholarly journals Tuning the tetraethylammonium sensitivity of potassium channel Kcv by subunit combination

2012 ◽  
Vol 139 (4) ◽  
pp. 295-304 ◽  
Author(s):  
Qiulin Tan ◽  
Brandon Ritzo ◽  
Kai Tian ◽  
Li-Qun Gu
Keyword(s):  
Potassium Channel ◽  
Binding Site ◽  
Specific Interaction ◽  
Site Directed Mutagenesis ◽  
Molecular Probe ◽  
K Channel ◽  
Optimal Conformation ◽  
Subunit Combination ◽  
Insight Into

Tetraethylammonium (TEA) is a potassium (K+) channel inhibitor that has been extensively used as a molecular probe to explore the structure of channels’ ion pathway. In this study, we identified that Leu70 of the virus-encoded potassium channel Kcv is a key amino acid that plays an important role in regulating the channel’s TEA sensitivity. Site-directed mutagenesis of Leu70 can change the TEA sensitivity by 1,000-fold from ∼100 µM to ∼100 mM. Because no compelling trends exist to explain this amino acid’s specific interaction with TEA, the role of Leu70 at the binding site is likely to ensure an optimal conformation of the extracellular mouth that confers high TEA affinity. We further assembled the subunits of mutant and wt-Kcv into a series of heterotetramers. The differences in these heterochannels suggest that all of the four subunits in a Kcv channel additively participate in the TEA binding, and each of the four residues at the binding site independently contributes an equal binding energy. We therefore can present a series of mutant/wild-type tetramer combinations that can probe TEA over three orders of magnitude in concentration. This study may give insight into the mechanism for the interaction between the potassium channel and its inhibitor.

scholarly journals Selective cadmium regulation mediated by a cooperative binding mechanism in CadR

2019 ◽  
Vol 116 (41) ◽  
pp. 20398-20403 ◽  
Author(s):  
Xichun Liu ◽  
Qingyuan Hu ◽  
Jinmei Yang ◽  
Shanqing Huang ◽  
Tianbiao Wei ◽  
...  
Keyword(s):  
Transcriptional Activity ◽  
Spectroscopic Studies ◽  
Cooperative Binding ◽  
Binding Mechanism ◽  
Cadmium Ion ◽  
Dna Binding Domains ◽  
Binding Domains ◽  
Optimal Conformation ◽  
Living Organisms ◽  
Insight Into

Detoxification of the highly toxic cadmium element is essential for the survival of living organisms. Pseudomonas putida CadR, a MerR family transcriptional regulator, has been reported to exhibit an ultraspecific response to the cadmium ion. Our crystallographic and spectroscopic studies reveal that the extra cadmium selectivity of CadR is mediated by the unexpected cooperation of thiolate-rich site I and histidine-rich site II. Cadmium binding in site I mediates the reorientation of protein domains and facilitates the assembly of site II. Subsequently, site II bridge-links 2 DNA binding domains through ligands His140/His145 in the C-terminal histidine-rich tail. With dynamic transit between 2 conformational states, this bridge could stabilize the regulator into an optimal conformation that is critical for enhancing the transcriptional activity of the cadmium detoxification system. Our results provide dynamic insight into how nature utilizes the unique cooperative binding mechanism in multisite proteins to recognize cadmium ions specifically.

scholarly journals A Critical HA1 Neutralizing Domain of H5N1 Influenza in an Optimal Conformation Induces Strong Cross-Protection

PLoS ONE ◽  
2013 ◽  
Vol 8 (1) ◽  
pp. e53568 ◽  
Author(s):  
Lanying Du ◽  
Guangyu Zhao ◽  
Shihui Sun ◽  
Xiujuan Zhang ◽  
Xiaojun Zhou ◽  
...  
Keyword(s):  
Cross Protection ◽  
H5n1 Influenza ◽  
Optimal Conformation
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