相关实验视频

Updated: Jun 6, 2025

Author Spotlight: Unveiling Oxidative Phosphorylation System Dynamics and Mitochondrial Roles in Health and Disease

Author Spotlight: Unveiling Oxidative Phosphorylation System Dynamics and Mitochondrial Roles in Health and Disease

Published on: May 3, 2024

从本地组织中净化和表征线粒体生物依赖的碳氧酶

Nicholas O Schneider¹, Martin St Maurice¹

¹Department of Biological Sciences, Marquette University, Milwaukee, WI, United States.

Methods in enzymology

|November 21, 2024

概括

研究人员开发了一种新的,具有成本效益的方法,以从猪肝中分离依赖生物的碳氧酶. 这种方法为药物发现和酶研究提供了高收益,克服了复合表达系统的挑战.

科学领域:

生物化学生物化学
酶学是一种酶学.
代谢途径代谢途径

背景情况:

生物素依赖性碳氧酶对新陈代谢至关重要,参与关键生物分子合成.
在获得足够数量的哺乳动物生物依碳酸酶以用于研究方面存在挑战.
在细菌或细胞培养系统中的重组表达产生了少量的这些酶.

研究的目的:

建立一个高效和廉价的协议,以隔离哺乳动物线粒体生物素依赖的碳氧酶.
为获得这些酶提供了难以复合表达的替代方法.
促进酶动力学,结构和药物发现的下游研究.

主要方法:

从新鲜的猪肝中隔离线粒体生物依赖的碳酸酶的协议恢复和优化.
使用的猪肝作为pyruvate carboxylase,propionyl-CoA carboxylase和3-methylcrotonyl-CoA carboxylase的来源.使用的猪肝作为pyruvate carboxylase,propionyl-CoA carboxylase和3-methylcrotonyl-CoA carboxylase的来源.
专注于实现高产量和有效地从细胞污染物中分离.

主要成果:

通过使用恢复的协议,成功分离了线粒体生物依赖的碳氧酶.
从猪肝中获得高产量,为重组系统提供了具有成本效益的替代方案.
该协议是可扩展的,可以在48小时内完成.

结论:

开发的协议提供了一种有效和经济的方法,用于获得哺乳动物线粒体生物依赖的碳氧酶.
这种方法克服了以前用于研究目的的酶净化限制.
纯化酶适用于动力学,结构学和药物发现研究.

关键词:

3 - 甲基克罗尼尔-CoA碳氧酶的使用.生物素是生物素中的一种.生物素依赖的酶生物素依赖的酶烯-CoA碳氧化酶是什么?酸盐碳氧化酶是什么?链状动素是一种链状动素.

更多相关视频

Author Spotlight: Two-Step Tag-Free Isolation of Mitochondria for Improved Protein Discovery and Quantification

Author Spotlight: Two-Step Tag-Free Isolation of Mitochondria for Improved Protein Discovery and Quantification

Published on: June 2, 2023

Analysis of Mitochondrial Respiratory Chain Complexes in Cultured Human Cells using Blue Native Polyacrylamide Gel Electrophoresis and Immunoblotting

Analysis of Mitochondrial Respiratory Chain Complexes in Cultured Human Cells using Blue Native Polyacrylamide Gel Electrophoresis and Immunoblotting

Published on: February 12, 2019

相关概念视频

Porin Insertion in the Outer Mitochondrial Membrane

Porin Insertion in the Outer Mitochondrial Membrane

Porins are beta-barrel proteins translocated to the mitochondrial outer membrane through the TOM complex into the intermembrane space. Porin precursors bind TIM chaperones within the intermembrane space and are guided to the Sorting and Assembly Machinery complex or SAM complex on the outer mitochondrial membrane.
Three models describe the assembly of porins by the SAM complex and their insertion into the outer membrane. Model 1 suggests that porins are assembled outside the SAM channel as the...

Translocation of Proteins into the Mitochondria

Translocation of Proteins into the Mitochondria

Mitochondrial precursors are translocated to the internal subcompartments via independent mechanisms involving distinct protein machineries called translocases.
Sorting of outer membrane proteins:
Mitochondrial outer membrane proteins are of two types: the transmembrane, beta-barrel porins, and the membrane-anchored, alpha-helical proteins. Beta-barrel porin precursors are translocated by the TOM complex and inserted into the outer mitochondrial membrane by the SAM complex. In contrast,...

Structure of Porins

Structure of Porins

Mitochondria, chloroplasts, and gram-negative bacteria have transmembrane, beta-barrel proteins called porins to mediate the free diffusion of ions and metabolites across the membrane. Mitochondrial porin precursors contain conserved amino acid sequences called beta signals at their C-terminal. Beta signals have a motif of PoXGXXHyXHy (Po-Polar, X-Any amino acid, G-Glycine, Hy-LargeHydrophobic), which are crucial for precursor recognition to initiate precursor assembly. Beta-barrel...