Biochemical and structural analyses suggest that plasminogen activators coevolved with their cognate protein substrates and inhibitors

Agnieszka Jendroszek; Jeppe B Madsen; Andrés Chana-Muñoz; Daniel M Dupont; Anni Christensen; Frank Panitz; Ernst-Martin Füchtbauer; Simon C Lovell; Jan K Jensen

doi:10.1074/jbc.RA118.005419

Biochemical and structural analyses suggest that plasminogen activators coevolved with their cognate protein substrates and inhibitors

Agnieszka Jendroszek, Jeppe B Madsen, Andrés Chana-Muñoz, Daniel M Dupont, Anni Christensen, Frank Panitz, Ernst-Martin Füchtbauer, Simon C Lovell, Jan K Jensen

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Abstract

Protein sequences of members of the plasminogen activation system are present throughout the entire vertebrate phylum. This important and well-described proteolytic cascade is governed by numerous protease-substrate and protease-inhibitor interactions whose conservation is crucial to maintaining unchanged protein function throughout evolution. The pressure to preserve protein-protein interactions may lead to either co-conservation or covariation of binding interfaces. Here, we combined covariation analysis and structure-based prediction to analyze the binding interfaces of urokinase (uPA):plasminogen activator inhibitor-1 (PAI-1) and uPA:plasminogen complexes. We detected correlated variation between the S3-pocket-lining residues of uPA and the P3 residue of both PAI-1 and plasminogen. These residues are known to form numerous polar interactions in the human uPA:PAI-1 Michaelis complex. To test the effect of mutations that correlate with each other and have occurred during mammalian diversification on protein-protein interactions, we produced uPA, PAI-1, and plasminogen from human and zebrafish to represent mammalian and nonmammalian orthologs. Using single amino acid point substitutions in these proteins, we found that the binding interfaces of uPA:plasminogen and uPA:PAI-1 may have coevolved to maintain tight interactions. Moreover, we conclude that although the interaction areas between protease-substrate and protease-inhibitor are shared, the two interactions are mechanistically different. Compared with a protease cleaving its natural substrate, the interaction between a protease and its inhibitor is more complex and involves a more fine-tuned mechanism. Understanding the effects of evolution on specific protein interactions may help further pharmacological interventions of the plasminogen activation system and other proteolytic systems.

Original language	English
Journal	Journal of Biological Chemistry
Volume	294
Issue	10
Pages (from-to)	3794-3805
Number of pages	12
ISSN	0021-9258
DOIs	https://doi.org/10.1074/jbc.RA118.005419
Publication status	Published - 8 Mar 2019

Keywords

Amino Acid Sequence
Animals
Evolution, Molecular
Humans
Models, Molecular
Plasminogen Activator Inhibitor 1/metabolism
Plasminogen Activators/antagonists & inhibitors
Protein Binding
Protein Conformation
Urokinase-Type Plasminogen Activator/metabolism

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10.1074/jbc.RA118.005419

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6416416/pdf/zbc3794.pdf

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Jendroszek, A., Madsen, J. B., Chana-Muñoz, A., Dupont, D. M., Christensen, A., Panitz, F., Füchtbauer, E.-M., Lovell, S. C., & Jensen, J. K. (2019). Biochemical and structural analyses suggest that plasminogen activators coevolved with their cognate protein substrates and inhibitors. Journal of Biological Chemistry, 294(10), 3794-3805. https://doi.org/10.1074/jbc.RA118.005419

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abstract = "Protein sequences of members of the plasminogen activation system are present throughout the entire vertebrate phylum. This important and well-described proteolytic cascade is governed by numerous protease-substrate and protease-inhibitor interactions whose conservation is crucial to maintaining unchanged protein function throughout evolution. The pressure to preserve protein-protein interactions may lead to either co-conservation or covariation of binding interfaces. Here, we combined covariation analysis and structure-based prediction to analyze the binding interfaces of urokinase (uPA):plasminogen activator inhibitor-1 (PAI-1) and uPA:plasminogen complexes. We detected correlated variation between the S3-pocket-lining residues of uPA and the P3 residue of both PAI-1 and plasminogen. These residues are known to form numerous polar interactions in the human uPA:PAI-1 Michaelis complex. To test the effect of mutations that correlate with each other and have occurred during mammalian diversification on protein-protein interactions, we produced uPA, PAI-1, and plasminogen from human and zebrafish to represent mammalian and nonmammalian orthologs. Using single amino acid point substitutions in these proteins, we found that the binding interfaces of uPA:plasminogen and uPA:PAI-1 may have coevolved to maintain tight interactions. Moreover, we conclude that although the interaction areas between protease-substrate and protease-inhibitor are shared, the two interactions are mechanistically different. Compared with a protease cleaving its natural substrate, the interaction between a protease and its inhibitor is more complex and involves a more fine-tuned mechanism. Understanding the effects of evolution on specific protein interactions may help further pharmacological interventions of the plasminogen activation system and other proteolytic systems.",

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author = "Agnieszka Jendroszek and Madsen, {Jeppe B} and Andr{\'e}s Chana-Mu{\~n}oz and Dupont, {Daniel M} and Anni Christensen and Frank Panitz and Ernst-Martin F{\"u}chtbauer and Lovell, {Simon C} and Jensen, {Jan K}",

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Jendroszek, A, Madsen, JB, Chana-Muñoz, A, Dupont, DM, Christensen, A, Panitz, F, Füchtbauer, E-M, Lovell, SC & Jensen, JK 2019, 'Biochemical and structural analyses suggest that plasminogen activators coevolved with their cognate protein substrates and inhibitors', Journal of Biological Chemistry, vol. 294, no. 10, pp. 3794-3805. https://doi.org/10.1074/jbc.RA118.005419

Biochemical and structural analyses suggest that plasminogen activators coevolved with their cognate protein substrates and inhibitors. / Jendroszek, Agnieszka; Madsen, Jeppe B; Chana-Muñoz, Andrés et al.
In: Journal of Biological Chemistry, Vol. 294, No. 10, 08.03.2019, p. 3794-3805.

Research output: Contribution to journal/Conference contribution in journal/Contribution to newspaper › Journal article › Research › peer-review

TY - JOUR

T1 - Biochemical and structural analyses suggest that plasminogen activators coevolved with their cognate protein substrates and inhibitors

AU - Jendroszek, Agnieszka

AU - Madsen, Jeppe B

AU - Chana-Muñoz, Andrés

AU - Dupont, Daniel M

AU - Christensen, Anni

AU - Panitz, Frank

AU - Füchtbauer, Ernst-Martin

AU - Lovell, Simon C

AU - Jensen, Jan K

PY - 2019/3/8

Y1 - 2019/3/8

N2 - Protein sequences of members of the plasminogen activation system are present throughout the entire vertebrate phylum. This important and well-described proteolytic cascade is governed by numerous protease-substrate and protease-inhibitor interactions whose conservation is crucial to maintaining unchanged protein function throughout evolution. The pressure to preserve protein-protein interactions may lead to either co-conservation or covariation of binding interfaces. Here, we combined covariation analysis and structure-based prediction to analyze the binding interfaces of urokinase (uPA):plasminogen activator inhibitor-1 (PAI-1) and uPA:plasminogen complexes. We detected correlated variation between the S3-pocket-lining residues of uPA and the P3 residue of both PAI-1 and plasminogen. These residues are known to form numerous polar interactions in the human uPA:PAI-1 Michaelis complex. To test the effect of mutations that correlate with each other and have occurred during mammalian diversification on protein-protein interactions, we produced uPA, PAI-1, and plasminogen from human and zebrafish to represent mammalian and nonmammalian orthologs. Using single amino acid point substitutions in these proteins, we found that the binding interfaces of uPA:plasminogen and uPA:PAI-1 may have coevolved to maintain tight interactions. Moreover, we conclude that although the interaction areas between protease-substrate and protease-inhibitor are shared, the two interactions are mechanistically different. Compared with a protease cleaving its natural substrate, the interaction between a protease and its inhibitor is more complex and involves a more fine-tuned mechanism. Understanding the effects of evolution on specific protein interactions may help further pharmacological interventions of the plasminogen activation system and other proteolytic systems.

AB - Protein sequences of members of the plasminogen activation system are present throughout the entire vertebrate phylum. This important and well-described proteolytic cascade is governed by numerous protease-substrate and protease-inhibitor interactions whose conservation is crucial to maintaining unchanged protein function throughout evolution. The pressure to preserve protein-protein interactions may lead to either co-conservation or covariation of binding interfaces. Here, we combined covariation analysis and structure-based prediction to analyze the binding interfaces of urokinase (uPA):plasminogen activator inhibitor-1 (PAI-1) and uPA:plasminogen complexes. We detected correlated variation between the S3-pocket-lining residues of uPA and the P3 residue of both PAI-1 and plasminogen. These residues are known to form numerous polar interactions in the human uPA:PAI-1 Michaelis complex. To test the effect of mutations that correlate with each other and have occurred during mammalian diversification on protein-protein interactions, we produced uPA, PAI-1, and plasminogen from human and zebrafish to represent mammalian and nonmammalian orthologs. Using single amino acid point substitutions in these proteins, we found that the binding interfaces of uPA:plasminogen and uPA:PAI-1 may have coevolved to maintain tight interactions. Moreover, we conclude that although the interaction areas between protease-substrate and protease-inhibitor are shared, the two interactions are mechanistically different. Compared with a protease cleaving its natural substrate, the interaction between a protease and its inhibitor is more complex and involves a more fine-tuned mechanism. Understanding the effects of evolution on specific protein interactions may help further pharmacological interventions of the plasminogen activation system and other proteolytic systems.

KW - Amino Acid Sequence

KW - Animals

KW - Evolution, Molecular

KW - Humans

KW - Models, Molecular

KW - Plasminogen Activator Inhibitor 1/metabolism

KW - Plasminogen Activators/antagonists & inhibitors

KW - Protein Binding

KW - Protein Conformation

KW - Urokinase-Type Plasminogen Activator/metabolism

UR - http://www.scopus.com/inward/record.url?scp=85062630785&partnerID=8YFLogxK

U2 - 10.1074/jbc.RA118.005419

DO - 10.1074/jbc.RA118.005419

M3 - Journal article

C2 - 30651349

SN - 0021-9258

VL - 294

SP - 3794

EP - 3805

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

IS - 10

ER -

Biochemical and structural analyses suggest that plasminogen activators coevolved with their cognate protein substrates and inhibitors

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