Dynamics of Protein Turnover, a Missing Dimension in Proteomics

Abstract

Functional genomic experiments frequently involve a comparison of the levels of gene expression between two or more genetic, developmental, or physiological states. Such comparisons can be carried out at either the RNA (transcriptome) or protein (proteome) level, but there is often a lack of congruence between parallel analyses using these two approaches. To fully interpret protein abundance data from proteomic experiments, it is necessary to understand the contributions made by the opposing processes of synthesis and degradation to the transition between the states compared. Thus, there is a need for reliable methods to determine the rates of turnover of individual proteins at amounts comparable to those obtained in proteomic experiments. Here, we show that stable isotope-labeled amino acids can be used to define the rate of breakdown of individual proteins by inspection of mass shifts in tryptic fragments. The approach has been applied to an analysis of abundant proteins in glucoselimited yeast cells grown in aerobic chemostat culture at steady state. The average rate of degradation of 50 proteins was 2.2%/h, although some proteins were turned over at imperceptible rates, and others had degradation rates of almost 10%/h. This range of values suggests that protein turnover is a significant missing dimension in proteomic experiments and needs to be considered when assessing protein abundance data and comparing it to the relative abundance of cognate mRNA species.

Analysis of proteome dynamics in the mouse brain

“…The constant renewal of the protein population is an energy-intensive process, yet it allows the cell to rapidly modulate protein levels in response to changes in the environment (5,6). Proper proteome dynamics are critical to normal development and maintenance of health (7,8). For example, the dysregulation of protein turnover has been implicated in the aging process (9), increased degradation of the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel is a primary cause of cystic fibrosis (10), and the inability to clear protein aggregates leads to pathogenic accumulations in Alzheimer's, Parkinson's, Creutzfeldt-Jakob, and other age-related diseases (11).…”

Quantitative proteomic analysis of primary neurons reveals diverse changes in synaptic protein content in fmr1 knockout mice

“…This enrichment profile likely reflects the protein turnover of neurons in culture. Several studies have used stable isotope labeling to measure protein turnover rates on a large scale (25,26). However, these studies suffered from throughput and sensitivity limitations.…”

Continuous culture – making a comeback?

“…Indeed, attempts to correlate protein levels between genetic, developmental or physiological states with global gene expression profiles often fail to show congruence (Delneri et al, 2001). Pratt et al (2002) addressed a fundamental problem, the analysis of the antagonistic process of protein turnover. This was undertaken by using stable-isotope-labelled amino acids and analysing the breakdown of individual proteins by tryptic digest mass shifts.…”