Deep Proteogenomics of a Photosynthetic Cyanobacterium

Cyanobacteria, the evolutionary ancestors of plant chloroplasts,
contribute substantially to the Earth’s biogeochemical cycles
and are of great interest for a sustainable economy. Knowledge of
protein expression is the key to understanding cyanobacterial metabolism;
however, proteome studies in cyanobacteria are limited and cover only
a fraction of the theoretical proteome. Here, we performed a comprehensive
proteogenomic analysis of the model cyanobacterium Synechocystis sp. PCC 6803 to characterize the expressed (phospho)­proteome, re-annotate
known and discover novel open reading frames (ORFs). By mapping extensive
shotgun mass spectrometry proteomics data onto a six-frame translation
of the Synechocystis genome, we refined the genomic
annotation of 64 ORFs, including eight completely novel ORFs. Our
study presents the largest reported (phospho)­proteome dataset for
a unicellular cyanobacterium, covering the expression of about 80%
of the theoretical proteome under various cultivation conditions,
such as nitrogen or carbon limitation. We report 568 phosphorylated
S/T/Y sites that are present on numerous regulatory proteins, including
the transcriptional regulators cyAbrB1 and cyAbrB2. We also catalogue
the proteins that have never been detected under laboratory conditions
and found that a large portion of them is plasmid-encoded. This dataset
will serve as a resource, providing dedicated information on growth
condition-dependent protein expression and phosphorylation.