A Single Amino Acid Switch Alters the Isoprene Donor Specificity in Ribosomally Synthesized and Post-Translationally Modified Peptide Prenyltransferases

Estrada, Paola; Morita, Maho; Hao, Yue; Schmidt, Eric W.; Nair, Satish K.

doi:10.1021/jacs.8b05187

Title: A Single Amino Acid Switch Alters the Isoprene Donor Specificity in Ribosomally Synthesized and Post-Translationally Modified Peptide Prenyltransferases

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Abstract

Mutation at a single amino acid alters the isoprene donor specificity of prenyltransferases involved in the modification of ribosomally synthesized and post-translationally modified peptides (RiPPs). Though most characterized RiPP prenyltransferases carry out the regiospecific transfer of C₅ dimethylallyl donor to the side chain atoms on macrocyclic acceptor substrates, the elucidation of the cyanobactin natural product piricyclamide 70005E1 identifies an O-geranyl modification on Tyr, a reaction with little prior biochemical precedence. Reconstitution and kinetic studies of the presumptive geranyltransferase PirF shows that the enzyme utilizes a C₁₀ donor, with no C₅ transferase activity. The crystal structure of PirF reveals a single amino acid difference in the vicinity of the isoprene-binding pocket, relative to the C₅ utilizing enzymes. Remarkably, only a single amino acid mutation is necessary to completely switch the donor specificity from a C₅ to a C₁₀ prenyltransferase, and vice versa. Lastly, we demonstrate that these enzymes may be used for the chemospecific attachment of C₅ or C₁₀ lipid groups on lanthipeptides, an unrelated class of RiPP natural products. Overall, these studies represent a rare example where prenyl donor specificity can be discretely altered, which expands the arsenal of synthetic biology tools for tuning biological activities of peptide natural products.

Authors:

Estrada, Paola ^[1]; Morita, Maho ^[2]; Hao, Yue ^[3];

^[2];

^[1]

Univ. of Illinois at Urbana-Champaign, IL (United States)
Univ. of Utah, Salt Lake City, UT (United States)
Univ. of Illinois at Urbana-Champaign, IL (United States); Amgen Pharmaceuticals, Cambridge, MA (United States)

Publication Date:: 2018-06-20

Research Org.:: Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)

Sponsoring Org.:: National Institutes of Health (NIH)

OSTI Identifier:: 1544874

Grant/Contract Number:: GM122521; GM102602; GM079038

Resource Type:: Accepted Manuscript

Journal Name:: Journal of the American Chemical Society

Additional Journal Information:: Journal Volume: 140; Journal Issue: 26; Journal ID: ISSN 0002-7863

Publisher:: American Chemical Society (ACS)

Country of Publication:: United States

Language:: ENGLISH

Subject:: 37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY; Peptides and proteins; monomers; crystal structure; chemical structure; chemical specificity

Citation Formats


                    Estrada, Paola, Morita, Maho, Hao, Yue, Schmidt, Eric W., and Nair, Satish K. A Single Amino Acid Switch Alters the Isoprene Donor Specificity in Ribosomally Synthesized and Post-Translationally Modified Peptide Prenyltransferases.  United States: N. p., 2018. 
Web.  doi:10.1021/jacs.8b05187.

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                    Estrada, Paola, Morita, Maho, Hao, Yue, Schmidt, Eric W., & Nair, Satish K. A Single Amino Acid Switch Alters the Isoprene Donor Specificity in Ribosomally Synthesized and Post-Translationally Modified Peptide Prenyltransferases.  United States.  https://doi.org/10.1021/jacs.8b05187

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                    Estrada, Paola, Morita, Maho, Hao, Yue, Schmidt, Eric W., and Nair, Satish K. Wed .  
"A Single Amino Acid Switch Alters the Isoprene Donor Specificity in Ribosomally Synthesized and Post-Translationally Modified Peptide Prenyltransferases".  United States.  https://doi.org/10.1021/jacs.8b05187.  https://www.osti.gov/servlets/purl/1544874.

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@article{osti_1544874,

  title        = {A Single Amino Acid Switch Alters the Isoprene Donor Specificity in Ribosomally Synthesized and Post-Translationally Modified Peptide Prenyltransferases},

  author       = {Estrada, Paola and Morita, Maho and Hao, Yue and Schmidt, Eric W. and Nair, Satish K.},

  abstractNote = {Mutation at a single amino acid alters the isoprene donor specificity of prenyltransferases involved in the modification of ribosomally synthesized and post-translationally modified peptides (RiPPs). Though most characterized RiPP prenyltransferases carry out the regiospecific transfer of C5 dimethylallyl donor to the side chain atoms on macrocyclic acceptor substrates, the elucidation of the cyanobactin natural product piricyclamide 70005E1 identifies an O-geranyl modification on Tyr, a reaction with little prior biochemical precedence. Reconstitution and kinetic studies of the presumptive geranyltransferase PirF shows that the enzyme utilizes a C10 donor, with no C5 transferase activity. The crystal structure of PirF reveals a single amino acid difference in the vicinity of the isoprene-binding pocket, relative to the C5 utilizing enzymes. Remarkably, only a single amino acid mutation is necessary to completely switch the donor specificity from a C5 to a C10 prenyltransferase, and vice versa. Lastly, we demonstrate that these enzymes may be used for the chemospecific attachment of C5 or C10 lipid groups on lanthipeptides, an unrelated class of RiPP natural products. Overall, these studies represent a rare example where prenyl donor specificity can be discretely altered, which expands the arsenal of synthetic biology tools for tuning biological activities of peptide natural products.},

  doi          = {10.1021/jacs.8b05187},

  journal      = {Journal of the American Chemical Society},

  number       = 26,

  volume       = 140,

  place        = {United States},

  year         = {2018},

  month        = {6}

}

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