相关实验视频

Updated: May 10, 2025

A New Toolkit for Evaluating Gene Functions using Conditional Cas9 Stabilization

A New Toolkit for Evaluating Gene Functions using Conditional Cas9 Stabilization

Published on: September 2, 2021

基于结构的CRISPR/Cas9蛋白质分类:一种机器学习方法来阐明Cas9带

Sita Sirisha Madugula, Vindi M Jayasinghe-Arachchige, Charlene Norgan Radler

bioRxiv : the preprint server for biology

|April 28, 2025

概括

我们开发了一种机器学习方法,用于在CRISPR/Cas9基因编辑中映射全性网络. 这确定了关键的残留物和一个"静电谷",以设计更具体的Cas9变体.

科学领域:

生物化学生物化学
分子生物学分子生物学
生物信息学是一种生物信息学.

背景情况:

克里斯普尔/卡斯9基因编辑依赖于全调节来获得特异性和稳定性.
了解这些机制对于开发精确的Cas9变体具有更少的非目标效应至关重要.

研究的目的:

引入一种基于结构的新型机器学习 (ML) 方法,用于在Cas9.9中识别远程全网络.
在Streptococcus pyogenes Cas9 (SpCas9) 中系统地绘制和完善这些网络,以了解其稳定性和特异性.

主要方法:

在所有可用的Cas9结构上训练了一个ML模型,以分析Cas9结构景观.
应用了两阶段的SHAP特征选择策略,使用Cα-Cα间残留距离来识别关键的氨酸-氨酸 (Lys-Arg) 残留对.
利用分子动力学 (MD) 模拟和突变分析来研究已识别的全osteric 网络和静电谷.

主要成果:

确定了28个关键的lys-arg残留对,它们调解了spcas9的域间通信,稳定性和特异性.
在Lys-Arg对中发现了一种具有明显稳定行为的等级性全网络.
揭示了对SpCas9结构完整性至关重要的静电谷,突变破坏了其DNA结合的形状和影响特异性的稳定性.

结论:

开发了一个新的ML框架,整合了结构分析和MD模拟,以了解Cas9异构.
确定了关键的全残留物和静电谷,为设计高保真Cas9变体提供了合理的策略.
该方法适用于其他酶,以了解全调节和蛋白质设计.

更多相关视频

Gene Digital Circuits Based on CRISPR-Cas Systems and Anti-CRISPR Proteins

Gene Digital Circuits Based on CRISPR-Cas Systems and Anti-CRISPR Proteins

Published on: October 18, 2022

Genome Editing in Mammalian Cell Lines using CRISPR-Cas

Genome Editing in Mammalian Cell Lines using CRISPR-Cas

Published on: April 11, 2019

相关概念视频

CRISPR

CRISPR

Genome editing technologies allow scientists to modify an organism’s DNA via the addition, removal, or rearrangement of genetic material at specific genomic locations. These types of techniques could potentially be used to cure genetic disorders such as hemophilia and sickle cell anemia. One popular and widely used DNA-editing research tool that could lead to safe and effective cures for genetic disorders is the CRISPR-Cas9 system. CRISPR-Cas9 stands for Clustered Regularly Interspaced...

CRISPR and crRNAs

CRISPR and crRNAs

Bacteria and archaea are susceptible to viral infections just like eukaryotes; therefore, they have developed a unique adaptive immune system to protect themselves. Clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins (CRISPR-Cas) are present in more than 45% of known bacteria and 90% of known archaea.
The CRISPR-Cas system stores a copy of foreign DNA in the host genome and uses it to identify the foreign DNA upon reinfection. CRISPR-Cas has three different...

Allosteric Proteins-ATCase

Allosteric Proteins-ATCase

Binding sites linkages can regulate a protein's function. For example, enzyme activity is often regulated through a feedback mechanism where the end product of the biochemical process serves as an inhibitor.
Aspartate transcarbamoylase (ATCase) is a cytosolic enzyme that catalyzes the condensation of L-aspartate and carbamoyl phosphate to N-carbamoyl-L-aspartate. This reaction is the first step in pyrimidine biosynthesis. UTP and CTP, the end products of the pyrimidine synthesis...

Conserved Binding Sites

Conserved Binding Sites

Many proteins’ biological role depends on their interactions with their ligands, small molecules that bind to specific locations on the protein known as ligand-binding sites. Ligand-binding sites are often conserved among homologous proteins as these sites are critical for protein function.
Binding sites are often located in large pockets, and if their location on a protein’s surface is unknown, it can be predicted using various approaches. The energetic method computationally...

Homologous Recombination

Homologous Recombination

The basic reaction of homologous recombination (HR) involves two chromatids that contain DNA sequences sharing a significant stretch of identity. One of these sequences uses a strand from another as a template to synthesize DNA in an enzyme-catalyzed reaction. The final product is a novel amalgamation of the two substrates. To ensure an accurate recombination of sequences, HR is restricted to the S and G2 phases of the cell cycle. At these stages, the DNA has been replicated already and the...

Ligand Binding and Linkage

Ligand Binding and Linkage

Allosteric proteins have more than one ligand binding site; the binding of a ligand to any of these sites influences the binding of ligands to the other sites. When a protein is allosteric, its binding sites are called coupled or linked. In the case of enzymes, the site that binds to the substrate is known as the active site and the other site is known as the regulatory site. When a ligand binds to the regulatory site, this leads to conformational changes in the protein that can influence...