DESCRIPTION (provided by applicant): Proteus vulgaris (P. vulgaris) is a major cause of urinary tract bacterial infections. P. vulgaris can grow on medical equipment, such as catheters, in an antibiotic-resistant state known as biofilms. In biofilms, bacteria are highly adhesive and are protected from antibiotics by a self-produced extracellular matrix of DNA, protein and polysaccharides. Therefore, it is critical to have a fundamental understanding of the pathways leading to this antibiotic-resistant state. Our long-term objective is to elucidate the molecular mechanism of the ribosome-dependent RNase family of toxins, which are important for biofilm formation. This family of toxins is thought to be activated during biofilm formation and deletion o these toxins leads to biofilm defects and increased sensitivity to antibiotics. Host inhibition of growth B (HigB) is a member of this family and was first identified in P. vulgaris. HigB cleaves ribosome-bound messenger RNA (mRNA) at adenosine-rich sequences. Recent evidence strongly suggests that the manner in which HigB recognizes mRNA is novel because there is no strict codon requirement (i.e. recognition of all three bases). To understand the differences that account for this possible novel mechanism, we will elucidate the molecular mechanism of HigB. We hypothesize that HigB recognizes mRNA differently from the traditional codon-dependent mRNA decoding factors such as tRNAs and release factors. For our first aim, we will biochemically identify HigB amino acids involved in mRNA recognition and degradation using a combination of biochemical techniques. In our second aim, we will use X-ray crystallography to elucidate the structural basis by which HigB can recognize more than one mRNA codon in the context of the ribosome. Our proposal will provide novel insights into the molecular mechanism by which proteins such as toxins recognize mRNA. Also, molecular knowledge of ribosome-dependent RNase toxins is required for any development of biofilm-inhibiting drugs and my research will provide such insight.

PUBLIC HEALTH RELEVANCE: Bacterial infections are becoming a dangerous threat in hospital settings due to bacterial antibiotic resistance. In this proposal, I study the molecular basis by which a protein likely involved with bacterial antibiotic evasion functions.