一个高效的方法,用于过度生产的DNA聚合酶从Pyrococcus furiosus使用优化自诱导系统在大肠杆菌埃舍里奇亚
Moch Irfan Hadi1, Fina Amreta Laksmi2, Helbert3
1Faculty of Science and Technology, UIN Sunan Ampel, Surabaya, Indonesia.
World journal of microbiology & biotechnology
|September 18, 2024
概括
在一个定义的介质中优化自诱导显著提高了Pyrococcus furiosus DNA聚合酶 (Pfupol) 的产量,增长了2.6倍. 这种具有成本效益的方法产生了高度纯净和活性的pfupol,用于生物技术应用.
科学领域:
- 生物技术是生物技术.
- 分子生物学分子生物学
- 酶工程是什么? 酶工程是什么?
背景情况:
- 愤怒的PyrococcusDNA聚合酶 (Pfupol) 是一种具有重要的生物技术需求的高保真性酶.
- 目前使用大肠杆菌T7促进体表达的生产方法不具有成本效益.
- 优化pfupol生产对于满足不断扩大的工业需求至关重要.
研究的目的:
- 开发一种优化且具有成本效益的自诱导工艺,用于复合Pfupol表达.
- 为了提高生物技术应用的活性pfupol的产量和纯度.
主要方法:
- 在定义的介质中开发了一种自诱导过程,用于Pfupol表达.
- 优化营养度 (葡萄糖,糖醇,乳糖) 以获得最大的蛋白质产量.
- 在特定条件下在5升生物反应器中扩大生产规模 (200rpm,3vm,10%的注射剂).
主要成果:
- 与以前的方法相比,实现了71mg/L纯Pfupol的生产产量,比以前的方法增加了2.6倍.
- 纯化的Pfupol具有1445个单位/毫克的高特异活性.
- 优化的自动诱导工艺被证明是经济可行的和实用的.
结论:
- 开发的自诱导策略显著提高了Pfupol的生产效率和产量.
- 这种优化过程为生物技术行业提供了可扩展和具有成本效益的高活性pfupol来源.
相关概念视频
01:32Replication in Prokaryotes
24.7K
DNA replication has three main steps: initiation, elongation, and termination. Replication in prokaryotes begins when initiator proteins bind to the single origin of replication (ori) on the cell's circular chromosome. Replication then proceeds around the entire circle of the chromosome in each direction from the two replication forks, resulting in two DNA molecules.
Many Proteins Work Together to Replicate the Chromosome
Replication is coordinated and carried out by a host of specialized...
Many Proteins Work Together to Replicate the Chromosome
Replication is coordinated and carried out by a host of specialized...
24.7K
01:31Proofreading
6.2K
Synthesis of new DNA molecules is carried out by the enzyme DNA polymerase, which adds nucleotides on the daughter strand complementary to the template DNA strand. DNA polymerase has a higher affinity to add the correct base and ensures fidelity during DNA replication. Furthermore, it exhibits proofreading activity during replication, using an exonuclease domain that cuts off incorrect nucleotides from the nascent DNA strand.
Errors During Replication are Corrected by the DNA Polymerase...
Errors During Replication are Corrected by the DNA Polymerase...
6.2K
01:32PCR
205.0K
Overview
205.0K
02:10Translesion DNA Polymerases
9.9K
Translesion (TLS) polymerases rescue stalled DNA polymerases at sites of damaged bases by replacing the replicative polymerase and installing a nucleotide across the damaged site. Doing so, TLS allows additional time for the cell to repair the damage before resuming regular DNA replication.
TLS polymerases are found in all three domains of life - archaea, bacteria, and eukaryotes. Of the different classes of TLS polymerases, members of the Y family are fitted with specialized structures that...
TLS polymerases are found in all three domains of life - archaea, bacteria, and eukaryotes. Of the different classes of TLS polymerases, members of the Y family are fitted with specialized structures that...
9.9K
03:01The Replisome
33.2K
DNA replication is carried out by a large complex of proteins that act in a coordinated matter to achieve high-fidelity DNA replication. Together this complex is known as the DNA replication machinery or the replisome.
The synthesis of the leading and lagging strands is a highly coordinated process. To explain this, the “Trombone model” was proposed by Bruce Alberts in 1980. The DNA loop formation starts when a primer is synthesized on the parent lagging strand. The loop grows with...
The synthesis of the leading and lagging strands is a highly coordinated process. To explain this, the “Trombone model” was proposed by Bruce Alberts in 1980. The DNA loop formation starts when a primer is synthesized on the parent lagging strand. The loop grows with...
33.2K
02:31Replication in Eukaryotes
170.4K
Overview
170.4K