European Biophysics Journal最新文献_第10页

AAV analysis by sedimentation velocity analytical ultracentrifugation: beyond empty and full capsids 沉降速度超离心分析AAV分析:超越空衣壳和满衣壳

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-04-10 DOI: 10.1007/s00249-023-01646-z

Alexander E. Yarawsky, Valeria Zai-Rose, Hazel M. Cunningham, John W. Burgner 2nd, Michael T. DeLion, Lake N. Paul

{"title":"AAV analysis by sedimentation velocity analytical ultracentrifugation: beyond empty and full capsids","authors":"Alexander E. Yarawsky, Valeria Zai-Rose, Hazel M. Cunningham, John W. Burgner 2nd, Michael T. DeLion, Lake N. Paul","doi":"10.1007/s00249-023-01646-z","DOIUrl":"10.1007/s00249-023-01646-z","url":null,"abstract":"<div><p>The recent surge of therapeutic interest in recombinant adeno-associated viral (AAV) vectors for targeted DNA delivery has brought analytical ultracentrifugation (AUC) into the spotlight. A major concern during formulation of AAV therapeutics is purity of the active species (DNA-containing capsid, or “filled capsids”). Insertion of DNA into AAV is not a highly efficient process; thus, a significant amount of empty and partial/intermediate AAV molecules may exist. Recent guidance from the FDA includes limiting the presence of empty AAV capsids and other impurities to reduce immunotoxicity. While chromatographic techniques (SEC, SEC–MALS, AEX) are often used for empty and full capsid quantitation due to the ease of accessibility and familiarity among most biochemists, the resolution and sensitivity attained by sedimentation velocity (SV-AUC) in the formulation buffer and purification buffers is unmatched. Approaches for using SV-AUC to determine the empty-to-full capsid ratio have already been discussed by others; however, in this report, we focus on the importance of characterizing other impurities, such as free DNA, partially filled capsids, and aggregates that are recognized as species of concern for immunotoxicity. We also demonstrate the usefulness of applying multiple analyses (e.g., c(<i>s</i>), g(<i>s</i>*), WDA) in confirming the presence of and determining the hydrodynamic parameters of these various species.</p></div>","PeriodicalId":548,"journal":{"name":"European Biophysics Journal","volume":"52 4-5","pages":"353 - 366"},"PeriodicalIF":2.0,"publicationDate":"2023-04-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4408856","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 7

A new UltraScan module for the characterization and quantification of analytical buoyant density equilibrium experiments to determine AAV capsid loading 一个新的UltraScan模块，用于分析浮力密度平衡实验的表征和定量，以确定AAV衣壳负载

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-04-04 DOI: 10.1007/s00249-023-01641-4

Alexey Savelyev, Emre H. Brookes, Amy Henrickson, Borries Demeler

{"title":"A new UltraScan module for the characterization and quantification of analytical buoyant density equilibrium experiments to determine AAV capsid loading","authors":"Alexey Savelyev, Emre H. Brookes, Amy Henrickson, Borries Demeler","doi":"10.1007/s00249-023-01641-4","DOIUrl":"10.1007/s00249-023-01641-4","url":null,"abstract":"<div><p>A method for characterizing and quantifying peaks formed in an analytical buoyant density equilibrium (ABDE) experiment is presented. An algorithm is derived to calculate the concentration of the density forming gradient material at every point in the cell, provided the rotor speed, temperature, meniscus position, bottom of the cell position, and the loading concentration, molar mass, and partial specific volume of the density gradient-forming material are known. In addition, a new peak fitting algorithm has been developed which allows the user to automatically quantify the peaks formed in terms of density, apparent partial specific volume, and relative abundance. The method is suitable for both ionic and non-ionic density forming materials and can be used with data generated from the UV optical system as well as the AVIV fluorescence optical system. These methods have been programmed in a new UltraScan-III module (<i>us_abde</i>). Examples are shown that demonstrate the application of the new module to adeno-associated viral vector preparations and proteins.</p></div>","PeriodicalId":548,"journal":{"name":"European Biophysics Journal","volume":"52 4-5","pages":"311 - 320"},"PeriodicalIF":2.0,"publicationDate":"2023-04-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s00249-023-01641-4.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4150899","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 5

Investigation of dynamic solution interactions between NET-1 and UNC-5B by multi-wavelength analytical ultracentrifugation NET-1与UNC-5B动态溶液相互作用的多波长超离心分析研究

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-03-20 DOI: 10.1007/s00249-023-01644-1

Haben Gabir, Monika Gupta, Markus Meier, Fabian Heide, Manuel Koch, Joerg Stetefeld, Borries Demeler

引用次数: 2

Correction: Abstracts 更正:摘要

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-03-18 DOI: 10.1007/s00249-023-01645-0

引用次数: 0

The effects of lithium on human red blood cells studied using optical spectroscopy and laser trap 利用光谱学和激光陷阱研究了锂对人体红细胞的影响

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-03-17 DOI: 10.1007/s00249-023-01643-2

Yashveer Singh, Aniket Chowdhury, Raktim Dasgupta, Shovan Kumar Majumder

{"title":"The effects of lithium on human red blood cells studied using optical spectroscopy and laser trap","authors":"Yashveer Singh, Aniket Chowdhury, Raktim Dasgupta, Shovan Kumar Majumder","doi":"10.1007/s00249-023-01643-2","DOIUrl":"10.1007/s00249-023-01643-2","url":null,"abstract":"<div><p>\u0000Lithium has been the treatment of choice for patients with bipolar disorder<b>.</b> However, lithium overdose happens more frequently since it has a very narrow therapeutic range in blood, necessitating investigation of its adverse effects on blood cells. The possible changes that lithium exposure may have on functional and morphological characteristics of human red blood cells (RBCs) have been studied ex vivo using single-cell Raman spectroscopy, optical trapping, and membrane fluorescent probe. The Raman spectroscopy was performed with excitation at 532 nm light, which also results in simultaneous photoreduction of intracellular hemoglobin (Hb). The level of photoreduction of lithium-exposed RBCs was observed to decline with lithium concentration, indicating irreversible oxygenation of intracellular Hb from lithium exposure. The lithium exposure may also have an effect on RBC membrane, which was investigated via optical stretching in a laser trap and the results suggest lower membrane fluidity for the lithium-exposed RBCs. The membrane fluidity of RBCs was further studied using the Prodan generalized polarization method and the results verify the reduction of membrane fluidity upon lithium exposure.</p></div>","PeriodicalId":548,"journal":{"name":"European Biophysics Journal","volume":"52 1-2","pages":"91 - 100"},"PeriodicalIF":2.0,"publicationDate":"2023-03-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s00249-023-01643-2.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4692662","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A spectral decomposition quality assessment tool for multi-wavelength AUC experiments with UltraScan 基于UltraScan的多波长AUC实验光谱分解质量评估工具

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-03-17 DOI: 10.1007/s00249-023-01640-5

Saeed Mortezazadeh, Borries Demeler

引用次数: 2

Experimental extraction of Young’s modulus of MCF-7 tissue using atomic force microscopy and the spherical contact models 利用原子力显微镜和球面接触模型提取MCF-7组织的杨氏模量

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-03-16 DOI: 10.1007/s00249-023-01642-3

Mahdi Mirzaluo, Fateme Fereiduni, Moein Taheri, Mehdi Modabberifar

引用次数: 2

Molecular dynamics study of collective water vibrations in a DNA hydration shell DNA水合壳中集体水振动的分子动力学研究

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-03-15 DOI: 10.1007/s00249-023-01638-z

Tetiana Bubon, Oleksii Zdorevskyi, Sergiy Perepelytsya

{"title":"Molecular dynamics study of collective water vibrations in a DNA hydration shell","authors":"Tetiana Bubon, Oleksii Zdorevskyi, Sergiy Perepelytsya","doi":"10.1007/s00249-023-01638-z","DOIUrl":"10.1007/s00249-023-01638-z","url":null,"abstract":"<div><p>The structure of DNA double helix is stabilized by water molecules and metal counterions that form the ion-hydration shell around the macromolecule. Understanding the role of the ion-hydration shell in the physical mechanisms of the biological functioning of DNA requires detailed studies of its structure and dynamics at the atomistic level. In the present work, the study of collective vibrations of water molecules around the DNA double helix was performed within the framework of classical all-atom molecular dynamics methods. Calculating the vibrational density of states, the vibrations of water molecules in the low-frequency spectra ranged from <span>(sim)</span>30 to <span>(sim)</span>300 <span>(hbox {cm}^{-1})</span> were analyzed for the case of different regions of the DNA double helix (minor groove, major groove, and phosphate groups). The analysis revealed significant differences in the collective vibrations behavior of water molecules in the DNA hydration shell, compared to the vibrations of bulk water. All low-frequency modes of the DNA ion-hydration shell are shifted by about 15–20 <span>(hbox {cm}^{-1})</span> towards higher frequencies, which is more significant for water molecules in the minor groove region of the double helix. The interactions of water molecules with the atoms of the macromolecule induce intensity decrease of the mode of hydrogen-bond symmetrical stretching near 150 <span>(hbox {cm}^{-1})</span>, leading to the disappearance of this mode in the DNA spectra. The obtained results can provide an interpretation of the experimental data for DNA low-frequency spectra and may be important for the understanding of the processes of indirect protein–nucleic recognition.</p></div>","PeriodicalId":548,"journal":{"name":"European Biophysics Journal","volume":"52 1-2","pages":"69 - 79"},"PeriodicalIF":2.0,"publicationDate":"2023-03-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4623498","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Generation of unfolded outer membrane protein ensembles defined by hydrodynamic properties 由流体力学性质定义的未折叠外膜蛋白集合的生成

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-03-11 DOI: 10.1007/s00249-023-01639-y

Taylor Devlin, Patrick J. Fleming, Nicole Loza, Karen G. Fleming

{"title":"Generation of unfolded outer membrane protein ensembles defined by hydrodynamic properties","authors":"Taylor Devlin, Patrick J. Fleming, Nicole Loza, Karen G. Fleming","doi":"10.1007/s00249-023-01639-y","DOIUrl":"10.1007/s00249-023-01639-y","url":null,"abstract":"<div><p>Outer membrane proteins (OMPs) must exist as an unfolded ensemble while interacting with a chaperone network in the periplasm of Gram-negative bacteria. Here, we developed a method to model unfolded OMP (uOMP) conformational ensembles using the experimental properties of two well-studied OMPs. The overall sizes and shapes of the unfolded ensembles in the absence of a denaturant were experimentally defined by measuring the sedimentation coefficient as a function of urea concentration. We used these data to model a full range of unfolded conformations by parameterizing a targeted coarse-grained simulation protocol. The ensemble members were further refined by short molecular dynamics simulations to reflect proper torsion angles. The final conformational ensembles have polymer properties different from unfolded soluble and intrinsically disordered proteins and reveal inherent differences in the unfolded states that necessitate further investigation. Building these uOMP ensembles advances the understanding of OMP biogenesis and provides essential information for interpreting structures of uOMP-chaperone complexes.</p></div>","PeriodicalId":548,"journal":{"name":"European Biophysics Journal","volume":"52 4-5","pages":"415 - 425"},"PeriodicalIF":2.0,"publicationDate":"2023-03-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4471980","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 1

SViMULATE: a computer program facilitating interactive, multi-mode simulation of analytical ultracentrifugation data 模拟:一个计算机程序，促进交互式，多模式模拟分析超离心数据

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-03-09 DOI: 10.1007/s00249-023-01637-0

Chad A. Brautigam

{"title":"SViMULATE: a computer program facilitating interactive, multi-mode simulation of analytical ultracentrifugation data","authors":"Chad A. Brautigam","doi":"10.1007/s00249-023-01637-0","DOIUrl":"10.1007/s00249-023-01637-0","url":null,"abstract":"<div><p>The ability to simulate sedimentation velocity (SV) analytical ultracentrifugation (AUC) experiments has proved to be a valuable tool for research planning, hypothesis testing, and pedagogy. Several options for SV data simulation exist, but they often lack interactivity and require up-front calculations on the part of the user. This work introduces SViMULATE, a program designed to make AUC experimental simulation quick, straightforward, and interactive. SViMULATE takes user-provided parameters and outputs simulated AUC data in a format suitable for subsequent analyses, if desired. The user is not burdened by the necessity to calculate hydrodynamic parameters for simulated macromolecules, as the program can compute these properties on the fly. It also frees the user of decisions regarding simulation stop time. SViMULATE features a graphical view of the species that are under simulation, and there is no limit on their number. Additionally, the program emulates data from different experimental modalities and data-acquisition systems, including the realistic simulation of noise for the absorbance optical system. The executable is available for immediate download.</p></div>","PeriodicalId":548,"journal":{"name":"European Biophysics Journal","volume":"52 4-5","pages":"293 - 302"},"PeriodicalIF":2.0,"publicationDate":"2023-03-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4395165","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 3