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相关概念视频

Conserved Binding Sites01:49

Conserved Binding Sites

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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...
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Cooperative Binding of Transcription Regulators02:13

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Transcriptional regulators bind to specific cis-regulatory sequences in the DNA to regulate gene transcription. These cis-regulatory sequences are very short, usually less than ten nucleotide pairs in length. The short length means that there is a high probability of the exact same sequence randomly occurring throughout the genome.  Since regulators can also bind to groups of similar sequences, this further increases the chances of random binding. Transcriptional regulators form...
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Single-Strand DNA Binding Proteins01:03

Single-Strand DNA Binding Proteins

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For successful DNA replication, the unwinding of double-stranded DNA must be accompanied by stabilization and protection of the separated single strands of the DNA. This crucial task is performed by single-strand DNA-binding (SSB) proteins. They bind to the DNA in a sequence-independent manner, which means that the nitrogenous bases of the DNA need not be present in a specific order for binding of SSB proteins to it. The binding of SSB proteins straightens single-stranded DNA (ssDNA) and makes...
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Ligand Binding Sites02:40

Ligand Binding Sites

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Proteins are dynamic macromolecules that carry out a wide variety of essential processes; however, the activities of most proteins depend on their interactions with other molecules or ions, known as ligands.
Protein-ligand interactions are quite specific; even though numerous potential ligands surround a cellular protein at any given time, only a particular ligand can bind to that protein. Moreover, a ligand binds only to a dedicated area on the surface of the protein, known as the...
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Conservative Site-specific Recombination and Phase Variation02:53

Conservative Site-specific Recombination and Phase Variation

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Because the DNA segments are cut and reorganized in a direction-specific manner, site-specific recombination has emerged as an efficient genetic engineering technique. Flippase and Cyclization recombinases or Flp and Cre, respectively, are two members of the tyrosine recombinase family derived from bacteriophages, that are used to mediate site-specific DNA insertions, deletions, and targeted expression of proteins in mammalian cell lines.
The recognition sites for Cre recombinase called LoxP...
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Cis-regulatory Sequences02:02

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Cis-regulatory sequences are short fragments of non-coding DNA that are present on the same chromosomes as the genes that they regulate. These fragments serve as binding sites for transcriptional regulators, proteins that are responsible for controlling gene transcription and differential gene expression across cell types in eukaryotes. Cis-regulatory sequences can be close to the gene of interest or thousands of bases away in the DNA sequence; however, those sequences that are further away are...
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Protein WISDOM: A Workbench for In silico De novo Design of BioMolecules
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对特定序列的DNA结合蛋白的计算设计.

Cameron J Glasscock1,2, Robert Pecoraro1,2,3, Ryan McHugh1,2

  • 1Department of Biochemistry, University of Washington, Seattle, WA, USA.

bioRxiv : the preprint server for biology
|October 4, 2023
PubMed
概括

研究人员开发了一种计算方法来设计新的DNA结合蛋白 (DBPs). 这些工程DBP可以针对特定的DNA序列,用于基因编辑和调节的应用.

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科学领域:

  • 分子生物学分子生物学
  • 生物技术是生物技术.
  • 蛋白质工程是指蛋白质工程.

背景情况:

  • 序列特定的DNA结合蛋白 (DBPs) 在生物过程和生物技术中至关重要.
  • 具有新奇特性的工程DBP对基因组编辑等应用非常感兴趣.
  • 针对任意DNA目标的DBP的计算设计仍然是一个重大挑战.

研究的目的:

  • 开发一种用于设计小,序列特定的DNA结合蛋白的计算方法.
  • 通过实验验证设计的DBP与特定的DNA标相对应.
  • 在蜂环境中评估设计的DBP的功能.

主要方法:

  • 使用计算方法来设计小型DBP,通过主要沟相互作用准特定的DNA序列.
  • 实验查用于生成和验证五个不同的DNA标的结合剂.
  • 进行了晶体结构分析,以确认DBP-目标DNA复杂几何.

主要成果:

  • 设计的DBP展示了多达6个基点位置的特定匹配计算模型.
  • 实现了对目标DNA序列的高结合亲和度,解离常数低至30-100nM.
  • 设计的DBP-target复合体的晶体结构验证了计算设计方法的准确性.
  • 工程 DBPs 成功调节了细菌 (大肠杆菌) 和哺乳动物细胞中的基因转录.

结论:

  • 开发的计算方法在设计特定序列的DBP方面取得了重大进展.
  • 这种方法为创建用于基因调节和编辑应用的小型,易于交付的DBP提供了一条途径.
  • 这些发现为定制设计的蛋白质在合成生物学和治疗学中的更广泛应用铺平了道路.