A genomic integration platform for heterologous cargo encapsulation in 1,2-propanediol utilization bacterial microcompartments

Nichols, Taylor M.; Kennedy, Nolan W.; Tullman-Ercek, Danielle

doi:10.1016/j.bej.2020.107496

Title: A genomic integration platform for heterologous cargo encapsulation in 1,2-propanediol utilization bacterial microcompartments

Journal Article · Tue Jan 14 00:00:00 EST 2020 · Biochemical Engineering Journal

DOI:https://doi.org/10.1016/j.bej.2020.107496· OSTI ID:1803709

Nichols, Taylor M. ^[1]; Kennedy, Nolan W. ^[1];

^[1]

Northwestern University, Evanston, IL (United States)

Bacterial microcompartments (MCPs) are protein structures that encapsulate specific metabolic pathways in bacteria. The 1,2-propanediol utilization (Pdu) MCP in Salmonella enterica serovar Typhimurium LT2 encapsulates the pathway for 1,2-propanediol degradation to sequester a toxic intermediate, enable cofactor recycling, and enhance pathway flux. The Pdu MCP has potential as an enclosed scaffold for metabolic engineering applications. To successfully use Pdu MCPs for this purpose, however, methods to enable and control heterologous cargo encapsulation are critical. To this end, we have developed a genomic expression platform for cargo encapsulation in Pdu MCPs. We integrated signal sequence-tagged fluorescent reporters into the pdu operon in place of native Pdu enzymes and evaluated the resulting expression and encapsulation levels. We found that fluorescent reporters were successfully co-encapsulated, with varying relative encapsulation levels achieved when using different integration locus and signal sequence combinations. We also observed that the native Pdu signal sequences mediated different encapsulation efficiencies independent of expression levels. This work establishes the genomic integration platform as a viable method for controlling cargo encapsulation, expanding the toolkit toward engineering the Pdu MCP as a tunable nanobioreactor.

View Accepted Manuscript (DOE)

View Accepted Manuscript (Publisher)

Cite

Export

Save

Research Organization:: Northwestern Univ., Evanston, IL (United States)

Sponsoring Organization:: USDOE Office of Science (SC); Army Research Office (ARO); National Science Foundation (NSF); National Institutes of Health (NIH); International Institute for Nanotechnology (IIN); Keck Foundation; State of Illinois

Grant/Contract Number:: SC0019337; W911NF-16-1-0169; DGE-1842165; T32GM008449; ECCS-1542205; DMR-1720139

OSTI ID:: 1803709

Alternate ID(s):: OSTI ID: 1703921

Journal Information:: Biochemical Engineering Journal, Vol. 156, Issue C; ISSN 1369-703X

Publisher:: ElsevierCopyright Statement

Country of Publication:: United States

Language:: English

References (50)

A systems-level model reveals that 1,2-Propanediol utilization microcompartments enhance pathway flux through intermediate sequestration Jakobson, Christopher M.; Tullman-Ercek, Danielle; Slininger, Marilyn F. PLOS Computational Biology, Vol. 13, Issue 5 https://doi.org/10.1371/journal.pcbi.1005525	journal	May 2017
Stochastic reaction–diffusion simulation of enzyme compartmentalization reveals improved catalytic efficiency for a synthetic metabolic pathway Conrado, Robert J.; Mansell, Thomas J.; Varner, Jeffrey D. Metabolic Engineering, Vol. 9, Issue 4 https://doi.org/10.1016/j.ymben.2007.05.002	journal	July 2007
Improved monomeric red, orange and yellow fluorescent proteins derived from Discosoma sp. red fluorescent protein Shaner, Nathan C.; Campbell, Robert E.; Steinbach, Paul A. Nature Biotechnology, Vol. 22, Issue 12 https://doi.org/10.1038/nbt1037	journal	November 2004
PduL Is an Evolutionarily Distinct Phosphotransacylase Involved in B12-Dependent 1,2-Propanediol Degradation by Salmonella enterica Serovar Typhimurium LT2 Liu, Y.; Leal, N. A.; Sampson, E. M. Journal of Bacteriology, Vol. 189, Issue 5 https://doi.org/10.1128/JB.01151-06	journal	December 2006
Salmonella typhimurium LT2 Catabolizes Propionate via the 2-Methylcitric Acid Cycle Horswill, Alexander R.; Escalante-Semerena, Jorge C. Journal of Bacteriology, Vol. 181, Issue 18 https://doi.org/10.1128/JB.181.18.5615-5623.1999	journal	September 1999
Microcompartments for B12-Dependent 1,2-Propanediol Degradation Provide Protection from DNA and Cellular Damage by a Reactive Metabolic Intermediate Sampson, E. M.; Bobik, T. A. Journal of Bacteriology, Vol. 190, Issue 8, p. 2966-2971 https://doi.org/10.1128/JB.01925-07	journal	February 2008
The Propanediol Utilization (pdu) Operon ofSalmonella enterica Serovar Typhimurium LT2 Includes Genes Necessary for Formation of Polyhedral Organelles Involved in Coenzyme B12-Dependent 1,2-Propanediol Degradation † Bobik, Thomas A.; Havemann, Gregory D.; Busch, Robert J. Journal of Bacteriology, Vol. 181, Issue 19 https://doi.org/10.1128/JB.181.19.5967-5975.1999	journal	January 1999
In vivo co-localization of enzymes on RNA scaffolds increases metabolic production in a geometrically dependent manner Sachdeva, Gairik; Garg, Abhishek; Godding, David Nucleic Acids Research, Vol. 42, Issue 14 https://doi.org/10.1093/nar/gku617	journal	July 2014
Genetic Analysis of the Protein Shell of the Microcompartments Involved in Coenzyme B12-Dependent 1,2-Propanediol Degradation by Salmonella Cheng, S.; Sinha, S.; Fan, C. Journal of Bacteriology, Vol. 193, Issue 6 https://doi.org/10.1128/JB.01473-10	journal	January 2011
PduP is a coenzyme-a-acylating propionaldehyde dehydrogenase associated with the polyhedral bodies involved in B 12 -dependent 1,2-propanediol degradation by Salmonella enterica serovar Typhimurium LT2 Leal, Nicole A.; Havemann, Gregory D.; Bobik, Thomas A. Archives of Microbiology, Vol. 180, Issue 5 https://doi.org/10.1007/s00203-003-0601-0	journal	November 2003
Golden Gate Shuffling: A One-Pot DNA Shuffling Method Based on Type IIs Restriction Enzymes Engler, Carola; Gruetzner, Ramona; Kandzia, Romy PLoS ONE, Vol. 4, Issue 5 https://doi.org/10.1371/journal.pone.0005553	journal	May 2009
The N-Terminal Region of the Medium Subunit (PduD) Packages Adenosylcobalamin-Dependent Diol Dehydratase (PduCDE) into the Pdu Microcompartment Fan, C.; Bobik, T. A. Journal of Bacteriology, Vol. 193, Issue 20 https://doi.org/10.1128/JB.05661-11	journal	August 2011
Use of modular, synthetic scaffolds for improved production of glucaric acid in engineered E. coli Moon, Tae Seok; Dueber, John E.; Shiue, Eric Metabolic Engineering, Vol. 12, Issue 3 https://doi.org/10.1016/j.ymben.2010.01.003	journal	May 2010
A rapid flow cytometry assay for the relative quantification of protein encapsulation into bacterial microcompartments Kim, Edward Y.; Tullman-Ercek, Danielle Biotechnology Journal, Vol. 9, Issue 3 https://doi.org/10.1002/biot.201300391	journal	January 2014
In vitro analysis of the interactions between the PocR regulatory protein and the promoter region of the cobalamin biosynthetic (cob) operon of Salmonella typhimurium LT2. Rondon, M. R.; Escalante-Semerena, J. C. Journal of bacteriology, Vol. 178, Issue 8 https://doi.org/10.1128/JB.178.8.2196-2203.1996	journal	January 1996
A Taxonomy of Bacterial Microcompartment Loci Constructed by a Novel Scoring Method Axen, Seth D.; Erbilgin, Onur; Kerfeld, Cheryl A. PLoS Computational Biology, Vol. 10, Issue 10 https://doi.org/10.1371/journal.pcbi.1003898	journal	October 2014
Short N-terminal sequences package proteins into bacterial microcompartments Fan, C.; Cheng, S.; Liu, Y. Proceedings of the National Academy of Sciences, Vol. 107, Issue 16 https://doi.org/10.1073/pnas.0913199107	journal	March 2010
Localization of Proteins to the 1,2-Propanediol Utilization Microcompartment by Non-native Signal Sequences Is Mediated by a Common Hydrophobic Motif Jakobson, Christopher M.; Kim, Edward Y.; Slininger, Marilyn F. Journal of Biological Chemistry, Vol. 290, Issue 40 https://doi.org/10.1074/jbc.M115.651919	journal	August 2015
The PduL Phosphotransacylase Is Used To Recycle Coenzyme A within the Pdu Microcompartment Liu, Yu; Jorda, Julien; Yeates, Todd O. Journal of Bacteriology, Vol. 197, Issue 14 https://doi.org/10.1128/JB.00056-15	journal	May 2015
Improved Production of l-Threonine in Escherichia coli by Use of a DNA Scaffold System Lee, Jun Hyoung; Jung, Suk-Chae; Bui, Le Minh Applied and Environmental Microbiology, Vol. 79, Issue 3 https://doi.org/10.1128/AEM.02578-12	journal	November 2012
Organization of Intracellular Reactions with Rationally Designed RNA Assemblies Delebecque, C. J.; Lindner, A. B.; Silver, P. A. Science, Vol. 333, Issue 6041 https://doi.org/10.1126/science.1206938	journal	June 2011
Bacterial microcompartments Kerfeld, Cheryl A.; Aussignargues, Clement; Zarzycki, Jan Nature Reviews Microbiology, Vol. 16, Issue 5 https://doi.org/10.1038/nrmicro.2018.10	journal	March 2018
The PduM Protein Is a Structural Component of the Microcompartments Involved in Coenzyme B12-Dependent 1,2-Propanediol Degradation by Salmonella enterica Sinha, S.; Cheng, S.; Fan, C. Journal of Bacteriology, Vol. 194, Issue 8 https://doi.org/10.1128/JB.06529-11	journal	February 2012
Synthetic protein scaffolds provide modular control over metabolic flux Dueber, John E.; Wu, Gabriel C.; Malmirchegini, G. Reza Nature Biotechnology, Vol. 27, Issue 8, p. 753-759 https://doi.org/10.1038/nbt.1557	journal	August 2009
Spatially organizing biochemistry: choosing a strategy to translate synthetic biology to the factory Jakobson, Christopher M.; Tullman-Ercek, Danielle; Mangan, Niall M. Scientific Reports, Vol. 8, Issue 1 https://doi.org/10.1038/s41598-018-26399-0	journal	May 2018
PduA Is a Shell Protein of Polyhedral Organelles Involved in Coenzyme B12-Dependent Degradation of 1,2-Propanediol in Salmonella enterica Serovar Typhimurium LT2 Havemann, G. D.; Sampson, E. M.; Bobik, T. A. Journal of Bacteriology, Vol. 184, Issue 5 https://doi.org/10.1128/JB.184.5.1253-1261.2002	journal	March 2002
NIH Image to ImageJ: 25 years of image analysis Schneider, Caroline A.; Rasband, Wayne S.; Eliceiri, Kevin W. Nature Methods, Vol. 9, Issue 7 https://doi.org/10.1038/nmeth.2089	journal	June 2012
De novo design of signal sequences to localize cargo to the 1,2-propanediol utilization microcompartment : Design of Signal Sequences Jakobson, Christopher M.; Slininger Lee, Marilyn F.; Tullman-Ercek, Danielle Protein Science, Vol. 26, Issue 5 https://doi.org/10.1002/pro.3144	journal	April 2017
De novo targeting to the cytoplasmic and luminal side of bacterial microcompartments Lee, Matthew J.; Mantell, Judith; Brown, Ian R. Nature Communications, Vol. 9, Issue 1 https://doi.org/10.1038/s41467-018-05922-x	journal	August 2018
Effect of Genomic Integration Location on Heterologous Protein Expression and Metabolic Engineering in E. coli Englaender, Jacob A.; Jones, J. Andrew; Cress, Brady F. ACS Synthetic Biology, Vol. 6, Issue 4 https://doi.org/10.1021/acssynbio.6b00350	journal	January 2017
Structural Insight into the Mechanisms of Transport across the Salmonella enterica Pdu Microcompartment Shell Crowley, Christopher S.; Cascio, Duilio; Sawaya, Michael R. Journal of Biological Chemistry, Vol. 285, Issue 48 https://doi.org/10.1074/jbc.M110.160580	journal	September 2010
Characterization of the PduS Cobalamin Reductase of Salmonella enterica and Its Role in the Pdu Microcompartment Cheng, S.; Bobik, T. A. Journal of Bacteriology, Vol. 192, Issue 19 https://doi.org/10.1128/JB.00575-10	journal	July 2010
Selective molecular transport through the protein shell of a bacterial microcompartment organelle Chowdhury, Chiranjit; Chun, Sunny; Pang, Allan Proceedings of the National Academy of Sciences, Vol. 112, Issue 10 https://doi.org/10.1073/pnas.1423672112	journal	February 2015
Optimal Compartmentalization Strategies for Metabolic Microcompartments Hinzpeter, Florian; Gerland, Ulrich; Tostevin, Filipe Biophysical Journal, Vol. 112, Issue 4 https://doi.org/10.1016/j.bpj.2016.11.3194	journal	February 2017
Cargo encapsulation in bacterial microcompartments: Methods and analysis Nichols, Taylor M.; Kennedy, Nolan W.; Tullman-Ercek, Danielle Methods in Enzymology https://doi.org/10.1016/bs.mie.2018.12.009	book	January 2019
A set of recombineering plasmids for gram-negative bacteria Datta, Simanti; Costantino, Nina; Court, Donald L. Gene, Vol. 379, p. 109-115 https://doi.org/10.1016/j.gene.2006.04.018	journal	September 2006
The poc locus is required for 1,2-propanediol-dependent transcription of the cobalamin biosynthetic (cob) and propanediol utilization (pdu) genes of Salmonella typhimurium. Rondon, M. R.; Escalante-Semerena, J. C. Journal of Bacteriology, Vol. 174, Issue 7 https://doi.org/10.1128/JB.174.7.2267-2272.1992	journal	January 1992
Tuning the Catalytic Activity of Subcellular Nanoreactors Jakobson, Christopher M.; Chen, Yiqun; Slininger, Marilyn F. Journal of Molecular Biology, Vol. 428, Issue 15 https://doi.org/10.1016/j.jmb.2016.07.006	journal	July 2016
FACS-optimized mutants of the green fluorescent protein (GFP) Cormack, Brendan P.; Valdivia, Raphael H.; Falkow, Stanley Gene, Vol. 173, Issue 1 https://doi.org/10.1016/0378-1119(95)00685-0	journal	January 1996
Cytoplasmic degradation of ssrA-tagged proteins: Cellular degradation of GFP-ssrA Farrell, Christopher M.; Grossman, Alan D.; Sauer, Robert T. Molecular Microbiology, Vol. 57, Issue 6 https://doi.org/10.1111/j.1365-2958.2005.04798.x	journal	August 2005
Solution Structure of a Bacterial Microcompartment Targeting Peptide and Its Application in the Construction of an Ethanol Bioreactor Lawrence, Andrew D.; Frank, Stefanie; Newnham, Sarah ACS Synthetic Biology, Vol. 3, Issue 7 https://doi.org/10.1021/sb4001118	journal	November 2013
Recombineering: Genetic Engineering in Bacteria Using Homologous Recombination Thomason, Lynn; Court, Donald L.; Bubunenko, Mikail Current Protocols in Molecular Biology, Vol. 78, Issue 1 https://doi.org/10.1002/0471142727.mb0116s78	journal	April 2007
DNA-guided assembly of biosynthetic pathways promotes improved catalytic efficiency Conrado, Robert J.; Wu, Gabriel C.; Boock, Jason T. Nucleic Acids Research, Vol. 40, Issue 4 https://doi.org/10.1093/nar/gkr888	journal	October 2011
The PduQ Enzyme Is an Alcohol Dehydrogenase Used to Recycle NAD+ Internally within the Pdu Microcompartment of Salmonella enterica Cheng, Shouqiang; Fan, Chenguang; Sinha, Sharmistha PLoS ONE, Vol. 7, Issue 10 https://doi.org/10.1371/journal.pone.0047144	journal	October 2012
Employing bacterial microcompartment technology to engineer a shell-free enzyme-aggregate for enhanced 1,2-propanediol production in Escherichia coli Lee, Matthew J.; Brown, Ian R.; Juodeikis, Rokas Metabolic Engineering, Vol. 36 https://doi.org/10.1016/j.ymben.2016.02.007	journal	July 2016
Rapid and Efficient One-Step Metabolic Pathway Integration in E. coli Bassalo, Marcelo C.; Garst, Andrew D.; Halweg-Edwards, Andrea L. ACS Synthetic Biology, Vol. 5, Issue 7 https://doi.org/10.1021/acssynbio.5b00187	journal	April 2016
Propionyl Coenzyme A Is a Common Intermediate in the 1,2-Propanediol and Propionate Catabolic Pathways Needed for Expression of the prpBCDE Operon during Growth of Salmonella enterica on 1,2-Propanediol Palacios, Sergio; Starai, Vincent J.; Escalante-Semerena, Jorge C. Journal of Bacteriology, Vol. 185, Issue 9 https://doi.org/10.1128/JB.185.9.2802-2810.2003	journal	May 2003
Synthetic Protein Scaffolds for Biosynthetic Pathway Colocalization on Lipid Droplet Membranes Lin, Jyun-Liang; Zhu, Jie; Wheeldon, Ian ACS Synthetic Biology, Vol. 6, Issue 8 https://doi.org/10.1021/acssynbio.7b00041	journal	May 2017
The ClpXP and ClpAP proteases degrade proteins with carboxy-terminal peptide tails added by the SsrA-tagging system Gottesman, S.; Roche, E.; Zhou, Y. Genes & Development, Vol. 12, Issue 9 https://doi.org/10.1101/gad.12.9.1338	journal	May 1998
Engineering bacterial microcompartments with heterologous enzyme cargos Wagner, Hanna J.; Capitain, Charlotte C.; Richter, Karsten Engineering in Life Sciences, Vol. 17, Issue 1 https://doi.org/10.1002/elsc.201600107	journal	September 2016

Similar Records

Linking the Salmonella enterica 1,2-Propanediol Utilization Bacterial Microcompartment Shell to the Enzymatic Core via the Shell Protein PduB

Journal Article · Mon May 16 00:00:00 EDT 2022 · Journal of Bacteriology · OSTI ID:1803709

Kennedy, Nolan W.; Mills, Carolyn E.; Abrahamson, Charlotte H.; +5 more

Vertex protein PduN tunes encapsulated pathway performance by dictating bacterial metabolosome morphology

Journal Article · Wed Jun 29 00:00:00 EDT 2022 · Nature Communications · OSTI ID:1803709

Mills, Carolyn E.; Waltmann, Curt; Archer, Andre G.; +9 more

Self-assembling Shell Proteins PduA and PduJ have Essential and Redundant Roles in Bacterial Microcompartment Assembly

Journal Article · Sat Nov 21 00:00:00 EST 2020 · Journal of Molecular Biology · OSTI ID:1803709

Kennedy, Nolan W.; Ikonomova, Svetlana P.; Slininger Lee, Marilyn; +2 more

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
bacterial microcompartments
Salmonella
genomic integration
cargo encapsulation
metabolic engineering
Pdu

Title: A genomic integration platform for heterologous cargo encapsulation in 1,2-propanediol utilization bacterial microcompartments

Citation Formats

References (50)

Similar Records

Related Subjects