Proteomic survey of the Streptomyces coelicolor nucleoid

Nucleoid-associated proteins (NAPs) are small, highly abundant transcriptional regulators with low sequence specificity which are involved in multiple DNA-related processes including gene expression, DNA protection, recombination/repair and nucleoid structuring. Through these functions they are able to regulate important phenotypic properties including virulence, secondary metabolism and stress resistance. However the set of NAPs known within the Actinobacteria is small and incomplete. The missing proteins are likely to be key regulators of irirulence in pathogens such as Mycobacterium tuberculosis and also of development and secondary metabolism in industrially-important species such as Streptomyces. Here, we use label-free LC MS/MS to systematically search for novel NAPs in isolated nucleoids of the model actinomycete Streptomyces coelicolor. Based on the criteria of high abundance (emPAI scord) and predicted DNA-binding ability (DNAbinder score) we identified a set of 24 proteins with a high predicted likelihood of being NAPs. The approach was deemed successful as the set included known major NAPs HupA, HupS, sIHF and Lsr2 as well as the global transcriptional regulators BldD and CRP and the pleiotropic response regulator AfsQ1. It also included a number of proteins whose functions are not yet known from recognisable classes of transcription factor (SCO2140, SC04493, SC01839, SC01210, SCO5405, SC04229, SCO3198) or from uncharacterised protein families (SCO5783, SCO5592, SCO3793, SC06482) which comprise a valuable set of candidates for further study. Biological significance In this paper we establish a robust protocol for preparing S. coelicolor nucleoids for mass spectrometric analysis and develop a workflow for identifying novel nucleoid-associated proteins (NAPs) by combining LC MS/MS with a bioinformatical analysis. The nucleoidassociated proteins of many species are known to be key regulators of virulence, stress tolerance and global patterns of gene expression. Identifying the missing nucleoid proteins of S. coelicolor is likely to have important implications for manipulating the production of secondary metabolites such as antibiotics. Candidate NAPs were identified. Several of these are highly conserved in clinically important species such as Mycobacterium and in many commercially important species such as Salinispora and Micromonospora which represent a vital source of novel drugs such as antibiotics, antifungals and anticancer agents. (C) 2013 Elsevier B.V. All rights reserved.