Abstract
Borrelia burgdorferi, the agent of Lyme disease, is a spirochetal pathogen with limited metabolic capabilities that survives under highly disparate host-specific conditions. However, the borrelial genome encodes several proteins of the mevalonate pathway (MP) that utilizes acetyl-CoA as a substrate leading to intermediate metabolites critical for biogenesis of peptidoglycan and post-translational modifications of proteins. In this study, we analyzed the MP and contributions of acetate in modulation of adaptive responses in B. burgdorferi. Reverse-transcription PCR revealed that components of the MP are transcribed as individual open reading frames. Immunoblot analysis using monospecific sera confirmed synthesis of members of the MP in B. burgdorferi. The rate-limiting step of the MP is mediated by HMG-CoA reductase (HMGR) via conversion of HMG-CoA to mevalonate. Recombinant borrelial HMGR exhibited a Km value of 132 µM with a Vmax of 1.94 µmol NADPH oxidized minute−1 (mg protein)−1 and was inhibited by statins. Total protein lysates from two different infectious, clonal isolates of B. burgdorferi grown under conditions that mimicked fed-ticks (pH 6.8/37°C) exhibited increased levels of HMGR while other members of the MP were elevated under unfed-tick (pH 7.6/23°C) conditions. Increased extra-cellular acetate gave rise to elevated levels of MP proteins along with RpoS, CsrABb and their respective regulons responsible for mediating vertebrate host-specific adaptation. Both lactone and acid forms of two different statins inhibited growth of B. burgdorferi strain B31, while overexpression of HMGR was able to partially overcome that inhibition. In summary, these studies on MP and contributions of acetate to host-specific adaptation have helped identify potential metabolic targets that can be manipulated to reduce the incidence of Lyme disease.
Highlights
Lyme disease is the most prevalent arthropod-borne disease in the United States with nearly 30,000 cases reported to the Centers for Disease Control and Prevention (CDC) in 2010 [1]
The presence of ORFs with sequence similarity to mevalonate kinase (Mvk; BB0688), phosphomevalonate kinase (Pmk; BB0687) and mevalonate decarboxylase (MvaD; BB0686) to homologs present in other bacteria and the conspicuous absence of members of the non-mevalonate pathway suggest that isopentenyl pyrophosphate, a central metabolic intermediate for cell wall biogenesis and posttranslational modification of proteins, is generated in B. burgdorferi via the MP
Effect of Statins on Growth of B. burgdorferi Based on the ability of select statins to inhibit recombinant borrelial hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase (HMGR), we evaluated the sensitivity of an infectious clonal isolate of B. burgdorferi strain B31-A3 with lovastatin and simvastatin either in the lactone or acid form prepared as described under Materials and Methods
Summary
Lyme disease is the most prevalent arthropod-borne disease in the United States with nearly 30,000 cases reported to the Centers for Disease Control and Prevention (CDC) in 2010 [1]. The incidence of Lyme disease is highest in geographic areas where there are increased ecological interactions between humans, reservoir hosts infected with B. burgdorferi, and transmission vectors [3,4,5,6,7,8,9,10,11]. Lyme disease is a multi-phasic disorder leading to severe arthritis, carditis, and/or neuroborreliosis if untreated during the early stages of infection [5,6,7,8,9,10]. While doxycycline is the antibiotic of choice for treatment during early stage Lyme disease, the efficacy of antibiotics can be variable in patients presenting with established, late-stage B. burgdorferi infection [12,13]. No vaccines are available against Lyme disease in humans [14], necessitating development of alternate strategies to limit the transmission of B. burgdorferi to humans and reducing the incidence of Lyme disease
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