Paired genomic and proteomic analysis of from waste sludge reveals an extensive repertoire of enzymes for breakdown of hydrocarbons.

Crude oil, primarily consisting of alkanes, and plastic derivatives are integral to modern society, yet their widespread use has led to persistent pollution that devastates ecosystems worldwide. Enzymatic and microbial biodegradation are highly desirable to become alternatives to conventional methods for the control of these pollutants in terrestrial and aquatic environments. Consequently, investigating the enzymatic machinery of specialized microbes that thrive in contaminated areas and possess the catabolic potential to degrade alkanes and plastics is of considerable interest. In this study, we present a genomic and proteomic analysis of RMS-02 that was isolated from sludge obtained from mixed food waste and sewage. Consistent with its ability to grow and persist in this environment, the genome of RMS-02 encodes enzymes to catabolize different types of polyphenols and aromatic compounds, including -cinnamic acid, -cresol, toluene, benzene, and benzoate. In addition, it encodes serine proteases, metallopeptidase, and transporters facilitating the uptake of amino acids and possibly small peptides, as well as enzymes potentially involved in the depolymerization of alkanes and 2-ketones. Proteomics analysis of RMS-02 revealed an extensive repertoire of enzymes involved in terminal and subterminal oxidation of medium- and long-chain alkanes and ketones, which were specifically more abundant during growth on a product that consists of a mixture of these compounds and an oxidized low-molecular-weight polyethylene (LMWPE). Substrate characterization following bacterial growth confirmed the selective utilization of alkanes with chain lengths of C10-C25 and 2-ketones of C13-C26, while the polymeric fraction of the substrate remained unaltered.IMPORTANCECrude oil and plastic pollution threaten ecosystems worldwide, creating an urgent need for sustainable remediation strategies. Microbial and enzymatic degradation provides a sustainable alternative to physical, chemical, and thermal treatments by biologically breaking down hydrocarbons into harmless products or recyclable monomers. Here, we describe a detailed genomic characterization of RMS-02, identifying core metabolic functions underlying its potential to utilize phenols, aromatic compounds, proteins, and small-chain hydrocarbons. Using proteomics, we show that a diverse enzymatic arsenal is deployed when growing on an oxidized LMWPE product that includes a mixture of alkanes and 2-ketones. Remarkably, the proteomics results were corroborated by advanced analysis of the spent substrate, confirming that RMS-02 metabolizes alkanes and 2-ketones but is unable to interact with the polymeric LMWPE component. Our results expand the understanding of the metabolic repertoire supporting 's survival and identify candidate enzymes with potential for the bioremediation of alkanes and 2-ketones.