Automated 16-Plex Plasma Proteomics with Real-Time Search and Ion Mobility Mass Spectrometry Enables Large-Scale Profiling in Naked Mole-Rats and Mice
Aleksandr Gaun, Kaitlyn N. Lewis Hardell, Niclas Olsson, Jonathon J. O’Brien, Sudha Gollapudi, Megan Smith, Graeme C. McAlister, Romain Huguet, Robert Keyser, Rochelle Buffenstein, Fiona E. McAllister
Abstract
Performing large-scale plasma proteome profiling is challenging due to limitations imposed by lengthy preparation and instrument time. We present a fully automated multiplexed proteome profiling platform (AutoMP3) using the Hamilton Vantage liquid handling robot capable of preparing hundreds to thousands of samples. To maximize protein depth in single-shot runs, we combined 16-plex Tandem Mass Tags (TMTpro) with high-field asymmetric waveform ion mobility spectrometry (FAIMS Pro) and real-time search (RTS). We quantified over 40 proteins/min/sample, doubling the previously published rates. We applied AutoMP3 to investigate the naked mole-rat plasma proteome both as a function of the circadian cycle and in response to ultraviolet (UV) treatment. In keeping with the lack of synchronized circadian rhythms in naked mole-rats, we find few circadian patterns in plasma proteins over the course of 48 h. Furthermore, we quantify many disparate changes between mice and naked mole-rats at both 48 h and one week after UV exposure. These species differences in plasma protein temporal responses could contribute to the pronounced cancer resistance observed in naked mole-rats. The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE [1] partner repository with the dataset identifier PXD022891.