Mesoporous Silica as Sorbents and Enzymatic Nanoreactors for Microbial Membrane Proteomics

The
membrane proteins of microbes are at the forefront of host
and parasite interactions. Having a general view of the functions
of microbial membrane proteins is vital for many biomedical studies
on microbiota. Nevertheless, due to the strong hydrophobicity and
low concentration of membrane proteins, it is hard to efficiently
enrich and digest the proteins for mass spectrometry analysis. Herein,
we design an enzymatic nanoreactor for the digestion of membrane proteins
using methylated well-ordered hexagonal mesoporous silica (Met-SBA-15).
The material can efficiently extract hydrophobic membrane proteins
and host the proteolysis in nanopores. The performance of the enzymatic
nanoreactor is first demonstrated using standard hydrophobic proteins
and then validated using membrane proteins extracted from Escherichia coli (E. coli) or a mixed bacterial
sample of eight strains. Using the nanoreactor, 431 membrane proteins
are identified from E. coli, accounting for 38.5%
of all membrane proteins of the species, which is much more than that
by the widely used in-solution digestion protocol. From the mixed
bacterial sample of eight strains, 1395 membrane proteins are identified
using the nanoreactor. On the contrary, the traditional in-solution
proteolysis workflow only leads to the identification of 477 membrane
proteins, demonstrating that the Met-SBA-15 can be offered as an excellent
tool for microbial membrane proteome research and is expected to be
used in human microbiota studies, e.g. host–microbe interactions.