由细菌毒素SsdA对序列上下文独立的单链DNA细胞因子去胺的结构基础
bioRxiv : the preprint server for biology
|September 16, 2024
概括
细菌脱氨酶毒素SsdA (单链DNA脱氨酶) 将单链DNA (ssDNA) 结合到独特的V形状,解释了其广泛的基质选择性. 这种酶是这种酶.
科学领域:
- 生物化学 生物化学
- 分子生物学分子生物学
- 结构生物学 结构生物学
背景情况:
- DNA除氨酶毒素调解细菌间的对抗性和细菌遗传多样性.
- 这些酶是基因组工程中的宝贵工具.
- 单链DNA脱氨酶 (SsdA) 是一种细菌脱氨酶毒素家族-2 (BaDTF2) 酶,与AID/APOBEC家族的酶不同,它可以降低低序列特异性的ssDNA细胞因子.
研究的目的:
- 为了确定SsdA与ssDNA基质复合的晶体结构.
- 阐明SsdA基质结合和广泛序列选择性的机制.
- 研究一种独特的β-氨基酸在SsdA的活性和稳定性中的作用.
主要方法:
- 进行X射线晶体学以获得SsdA-ssDNA复杂结构.
- 结构功能研究分析基质结合和酶活性.
- 局部定向突变发生,以创建和测试SsdA变体.
主要成果:
- 晶体结构揭示了一种新的V形ssDNA结合模式,芳香残留物使DNA在目标细胞酸中急剧曲.
- 结合模式解释了SsdA的广泛基质选择性,这是由于与侧面基的有限的序列特定接触造成的.
- 发现SsdA含有β-氨基酸异酸,对酶活性和毒素稳定性至关重要.
结论:
- 独特的V形ssDNA结合机制是SsdA广泛基质特异性的基础.
- β-氨基酸异酸对SsdA的功能和稳定性至关重要.
- 在人类细胞中活跃的工程SsdA突变体显示出未来基因组工程应用的潜力.
相关概念视频
Single-Strand DNA Binding Proteins
14.0K
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...
14.0K
Maxam-Gilbert Sequencing
11.1K
In the same year as the discovery of the Sanger sequencing method, another group of scientists, Allan Maxam and Walter Gilbert, demonstrated their chemical-cleavage method for DNA sequencing. The Maxam-Gilbert method relies on using different chemicals that can cleave the DNA sequence at specific sites, the separation of resulting DNA fragments of variable size using electrophoresis, and deciphering the DNA sequence from the resulting gel bands.
Challenges of the Maxam-Gilbert Method
The...
Challenges of the Maxam-Gilbert Method
The...
11.1K
Fixing Double-strand Breaks
12.5K
The double-stranded structure of DNA has two major advantages. First, it serves as a safe repository of genetic information where one strand serves as the back-up in case the other strand is damaged. Second, the double-helical structure can be wrapped around proteins called histones to form nucleosomes, which can then be tightly wound to form chromosomes. This way, DNA chains up to 2 inches long can be contained within microscopic structures in a cell. A double-stranded break not only damages...
12.5K
Overview of DNA Repair
30.9K
In order to be passed through generations, genomic DNA must be undamaged and error-free. However, every day, DNA in a cell undergoes several thousand to a million damaging events by natural causes and external factors. Ionizing radiation such as UV rays, free radicals produced during cellular respiration, and hydrolytic damage from metabolic reactions can alter the structure of DNA. Damages caused include single-base alteration, base dimerization, chain breaks, and cross-linkage.
Chemically...
Chemically...
30.9K
Base Excision Repair
22.2K
One of the common DNA damages is the chemical alteration of single bases by alkylation, oxidation, or deamination. The altered bases cause mispairing and strand breakage during replication. This type of damage causes minimal change to the DNA double helix structure and can be repaired by the base excision repair (BER) pathways. BER corrects damaged DNA sequences by removing the damaged base and restoring the original base sequence using the complementary strand as a template.
The first step of...
The first step of...
22.2K
Sanger Sequencing
753.8K
DNA sequencing is a fundamental technique that is routinely used in the biological sciences. This method can be applied to a range of questions at different scales - from the sequencing of a cloned DNA fragment or the study of a mutation in a gene up to whole-genome sequencing. However, despite the widespread use of sequencing today, it was not until 1977 that Fredrick Sanger and his collaborators developed the chain-termination method to decode DNA sequences. It relies on the separation of a...
753.8K


