2 research outputs found
Measuring H<sub>2</sub><sup>18</sup>O Tracer Incorporation on a QQQ-MS Platform Provides a Rapid, Transferable Screening Tool for Relative Protein Synthesis
Intracellular proteins are in a state of flux, continually
being
degraded into amino acids and resynthesized into new proteins. The
rate of this biochemical recycling process varies across proteins
and is emerging as an important consideration in drug discovery and
development. Here, we developed a triple-stage quadrupole mass spectrometry
assay based on product ion measurements at unit resolution and H<sub>2</sub><sup>18</sup>O stable tracer incorporation to measure relative
protein synthesis rates. As proof of concept, we selected to measure
the relative in vivo synthesis rate of ApoB100, an apolipoprotein
where elevated levels are associated with an increased risk of coronary
heart disease, in plasma-isolated very low density lipoprotein (VLDL)
and low density lipoprotein (LDL) in a mouse in vivo model. In addition,
serial time points were acquired to measure the relative in vivo synthesis
rate of mouse LDL ApoB100 in response to vehicle, microsomal triacylglycerol
transfer protein (MTP) inhibitor, and site-1 protease inhibitor, two
potential therapeutic targets to reduce plasma ApoB100 levels at 2
and 6 h post-tracer-injection. The combination of H<sub>2</sub><sup>18</sup>O tracer with the triple quadrupole mass spectrometry platform
creates an assay that is relatively quick and inexpensive to transfer
across different biological model systems, serving as an ideal rapid
screening tool for relative protein synthesis in response to treatment
Measuring H<sub>2</sub><sup>18</sup>O Tracer Incorporation on a QQQ-MS Platform Provides a Rapid, Transferable Screening Tool for Relative Protein Synthesis
Intracellular proteins are in a state of flux, continually
being
degraded into amino acids and resynthesized into new proteins. The
rate of this biochemical recycling process varies across proteins
and is emerging as an important consideration in drug discovery and
development. Here, we developed a triple-stage quadrupole mass spectrometry
assay based on product ion measurements at unit resolution and H<sub>2</sub><sup>18</sup>O stable tracer incorporation to measure relative
protein synthesis rates. As proof of concept, we selected to measure
the relative in vivo synthesis rate of ApoB100, an apolipoprotein
where elevated levels are associated with an increased risk of coronary
heart disease, in plasma-isolated very low density lipoprotein (VLDL)
and low density lipoprotein (LDL) in a mouse in vivo model. In addition,
serial time points were acquired to measure the relative in vivo synthesis
rate of mouse LDL ApoB100 in response to vehicle, microsomal triacylglycerol
transfer protein (MTP) inhibitor, and site-1 protease inhibitor, two
potential therapeutic targets to reduce plasma ApoB100 levels at 2
and 6 h post-tracer-injection. The combination of H<sub>2</sub><sup>18</sup>O tracer with the triple quadrupole mass spectrometry platform
creates an assay that is relatively quick and inexpensive to transfer
across different biological model systems, serving as an ideal rapid
screening tool for relative protein synthesis in response to treatment