Bacterial microcompartments (MCPs) are giant protein assemblies that serve as metabolic organelles in diverse bacteria found throughout the microbial world. These extraordinary structures are composed of thousands of subunits that assemble to form a polyhedral outer shell encapsulating a series of sequentially acting metabolic enzymes. MCPs typically encapsulate pathways that produce volatile or toxic intermediates that must be confined and metabolized to other compounds before diffusing out of the MCP and into the bacterial cytosol. MCPs confer special growth advantages to enteric bacteria and are linked to bacterial pathogenesis and the dissemination of enteric pathogens. Prior studies have focused primarily on selected MCP types, and important advances have been made. However, a number of mechanistic questions remain unanswered and some important MCP types are essentially uncharacterized. Prior work by our dual-PI team (Bobik and Yeates) focused primarily on the propanediol utilization (Pdu) MCP, which is used by Salmonella and other enteric bacteria to degrade 1,2-propanediol while sequestering a toxic intermediate, propionaldehyde. Our research in the previous cycle led to numerous important discoveries and critical insights into mechanistic aspects of how the Pdu MCP functions. Our key findings cover biological phenomena related to protein structure and assembly, molecular recognition, molecular transport, and molecular evolution. Our current proposal focuses on (1) remaining questions about the assembly and operation of the Pdu MCP of Salmonella, and (2) early-stage investigations into a new and diverse class of MCPs (which we identified bioinformatically) whose key internalized enzymes catalyze glycyl-radical-based reactions. Our continuing work on the Pdu MCP will answer outstanding questions about protein-protein interactions used to guide the assembly of the Pdu MCP? our earlier work led to the discovery of peptide targeting sequences that direct enzyme encapsulation by binding the interior surface of MCP shells. However, further experiments are required to paint a clearer picture about preferential associations by varied targeting sequences and their contribution to higher-order structural organization. In prior work, we also showed that pores through the shell proteins in the Pdu MCP have evolved for selective diffusive molecular transport of small molecules. In our continuing work, we propose experiments to investigate the dynamics and regulation of protein conformational changes that affect pore opening and closing in MCP shell proteins. The second part of the proposal focuses on the newly-defined and little-studied class of MCPs that encapsulate metabolic pathways dependent on glycyl-radical (Gr) enzymes. A number of Gr-MCPs are found in bacteria that inhabit the large intestine and which can infect the urinary tract. We will undertake work on three proposed Gr subtypes: one type that metabolizes 1,2- PD (similarly to the Pdu system but using unrelated enzymes), and two distinct subtypes that are believed to metabolize choline. Our new research on the Gr systems will lay the foundations for understanding their unique structures and mechanisms. We will answer questions about their composition, metabolic function, organization, and structure. As with our studies of the Pdu MCP, our interdisciplinary work will be guided by structural and genetic studies, especially of diverse shell proteins and the properties of their pores, which are at the heart of molecular transport phenomena in these systems. The Pdu and Gr systems will also be compared and contrasted to gain insights into the principles that underlie functional diversification of bacterial MCPs.

Public Health Relevance

Bacterial microcompartments (MCPs) are extraordinary but still poorly understood protein assemblies used by hundreds of bacterial species to sequester key cellular reactions within the confines of a protein shell or capsid; this protects the bacterial cytosol from the damaging effects of toxic intermediates produced by the reaction pathway. Our earlier work revealed many exciting discoveries, but much remains to be learned about these systems ? how they evolved, how their protein components are organized, and how the movement of substrate and product molecules in and out of the MCP is controlled. MCPs have important functions in many bacteria that inhabit or infect human hosts. Some of their activities appear to be important in bacterial pathogenesis and the dissemination of enteric pathogens. A deeper understanding of these poorly recognized biological systems will provide fundamentally new and useful insights into bacterial metabolism and evolution that may be helpful for control of pathogenic bacteria.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI081146-09
Application #
9419268
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Alexander, William A
Project Start
2009-07-17
Project End
2022-04-30
Budget Start
2018-05-01
Budget End
2019-04-30
Support Year
9
Fiscal Year
2018
Total Cost
Indirect Cost
Name
Iowa State University
Department
Biochemistry
Type
Earth Sciences/Resources
DUNS #
005309844
City
Ames
State
IA
Country
United States
Zip Code
50011
Lehman, Brent P; Chowdhury, Chiranjit; Bobik, Thomas A (2017) The N Terminus of the PduB Protein Binds the Protein Shell of the Pdu Microcompartment to Its Enzymatic Core. J Bacteriol 199:
Gan, Qinglei; Lehman, Brent P; Bobik, Thomas A et al. (2016) Expanding the genetic code of Salmonella with non-canonical amino acids. Sci Rep 6:39920
Jorda, J; Leibly, D J; Thompson, M C et al. (2016) Structure of a novel 13 nm dodecahedral nanocage assembled from a redesigned bacterial microcompartment shell protein. Chem Commun (Camb) 52:5041-4
Wheatley, Nicole M; Eden, Kevin D; Ngo, Joanna et al. (2016) A PII-Like Protein Regulated by Bicarbonate: Structural and Biochemical Studies of the Carboxysome-Associated CPII Protein. J Mol Biol 428:4013-4030
Chowdhury, Chiranjit; Chun, Sunny; Sawaya, Michael R et al. (2016) The function of the PduJ microcompartment shell protein is determined by the genomic position of its encoding gene. Mol Microbiol 101:770-83
Thompson, Michael C; Cascio, Duilio; Leibly, David J et al. (2015) An allosteric model for control of pore opening by substrate binding in the EutL microcompartment shell protein. Protein Sci 24:956-75
Jorda, Julien; Liu, Yu; Bobik, Thomas A et al. (2015) Exploring bacterial organelle interactomes: a model of the protein-protein interaction network in the Pdu microcompartment. PLoS Comput Biol 11:e1004067
Chowdhury, Chiranjit; Chun, Sunny; Pang, Allan et al. (2015) Selective molecular transport through the protein shell of a bacterial microcompartment organelle. Proc Natl Acad Sci U S A 112:2990-5
Liu, Yu; Jorda, Julien; Yeates, Todd O et al. (2015) The PduL Phosphotransacylase Is Used To Recycle Coenzyme A within the Pdu Microcompartment. J Bacteriol 197:2392-9
Sturms, Ryan; Streauslin, Nicholas A; Cheng, Shouqiang et al. (2015) In Salmonella enterica, Ethanolamine Utilization Is Repressed by 1,2-Propanediol To Prevent Detrimental Mixing of Components of Two Different Bacterial Microcompartments. J Bacteriol 197:2412-21

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