Development of an Enhanced Metaproteomic Approach for Deepening the Microbiome Characterization of the Human Infant Gut

The establishment of early life microbiota
in the human infant
gut is highly variable and plays a crucial role in host nutrient availability/uptake
and maturation of immunity. Although high-performance mass spectrometry
(MS)-based metaproteomics is a powerful method for the functional
characterization of complex microbial communities, the acquisition
of comprehensive metaproteomic information in human fecal samples
is inhibited by the presence of abundant human proteins. To alleviate
this restriction, we have designed a novel metaproteomic strategy
based on double filtering (DF) the raw samples, a method that fractionates
microbial from human cells to enhance microbial protein identification
and characterization in complex fecal samples from healthy premature
infants. This method dramatically improved the overall depth of infant
gut proteome measurement, with an increase in the number of identified
low-abundance proteins and a greater than 2-fold improvement in microbial
protein identification and quantification. This enhancement of proteome
measurement depth enabled a more extensive microbiome comparison between
infants by not only increasing the confidence of identified microbial
functional categories but also revealing previously undetected categories.