Conformational Dynamics and Catalytic Backups in a Hyper-thermostable Engineered Archaeal Protein Tyrosine Phosphatase
(2026) In JACS Au 6(1). p.59-81- Abstract
Protein tyrosine phosphatases (PTPs) are a family of enzymes that play important roles in regulating cellular signaling pathways. The activity of these enzymes is regulated by the motion of a catalytic loop that places a critical conserved aspartic acid side chain into the active site for acid–base catalysis upon loop closure. These enzymes also have a conserved phosphate-binding loop that is typically highly rigid and forms a well-defined anion-binding nest. The intimate links between loop dynamics and chemistry in these enzymes make PTPs an excellent model system for understanding the role of loop dynamics in protein function and evolution. In this context, archaeal PTPs, which have often evolved in extremophilic organisms, are highly... (More)
Protein tyrosine phosphatases (PTPs) are a family of enzymes that play important roles in regulating cellular signaling pathways. The activity of these enzymes is regulated by the motion of a catalytic loop that places a critical conserved aspartic acid side chain into the active site for acid–base catalysis upon loop closure. These enzymes also have a conserved phosphate-binding loop that is typically highly rigid and forms a well-defined anion-binding nest. The intimate links between loop dynamics and chemistry in these enzymes make PTPs an excellent model system for understanding the role of loop dynamics in protein function and evolution. In this context, archaeal PTPs, which have often evolved in extremophilic organisms, are highly understudied, despite their unusual biophysical properties. We present here an engineered chimeric PTP (ShufPTP) generated by shuffling the amino acid sequence of five extant hyperthermophilic archaeal PTPs. Despite ShufPTP’s high sequence similarity to its natural counterparts, it presents a suite of unique properties, including high flexibility of the phosphate binding P-loop, facile oxidation of the active-site cysteine, mechanistic promiscuity, and, most notably, hyperthermostability, with a denaturation temperature likely >130 °C (>8 °C higher than the highest recorded growth temperature of any archaeal strain). Our combined structural, biochemical, biophysical, and computational analysis provides insight both into how small steps in evolutionary space can radically modulate the biophysical properties of an enzyme and showcases the tremendous potential of archaeal enzymes for biotechnology, to generate novel enzymes capable of operating under extreme conditions.
(Less)
- author
- Yehorova, Dariia
; Alansson, Nikolas
; Shen, Ruidan
; Denson, Joshua M.
; Robinson, Michael
; Risso, Valeria A.
; Ramirez Molina, Nuria
; Loria, J. Patrick
; Gaucher, Eric A.
; Sanchez-Ruiz, Jose M.
; Hengge, Alvan C.
; Johnson, Sean J.
and Kamerlin, Shina C.L.
LU
(Less) - organization
- publishing date
- 2026-01
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- archaea, catalytic backups, protein conformational dynamics, protein tyrosine phosphatase, sequence shuffling
- in
- JACS Au
- volume
- 6
- issue
- 1
- pages
- 23 pages
- publisher
- The American Chemical Society (ACS)
- external identifiers
-
- pmid:41614192
- scopus:105028323925
- ISSN
- 2691-3704
- DOI
- 10.1021/jacsau.5c00756
- language
- English
- LU publication?
- yes
- id
- 137b92f8-1582-442b-80a2-18d50ff119bf
- date added to LUP
- 2026-02-19 14:43:52
- date last changed
- 2026-02-20 03:57:07
@article{137b92f8-1582-442b-80a2-18d50ff119bf,
abstract = {{<p>Protein tyrosine phosphatases (PTPs) are a family of enzymes that play important roles in regulating cellular signaling pathways. The activity of these enzymes is regulated by the motion of a catalytic loop that places a critical conserved aspartic acid side chain into the active site for acid–base catalysis upon loop closure. These enzymes also have a conserved phosphate-binding loop that is typically highly rigid and forms a well-defined anion-binding nest. The intimate links between loop dynamics and chemistry in these enzymes make PTPs an excellent model system for understanding the role of loop dynamics in protein function and evolution. In this context, archaeal PTPs, which have often evolved in extremophilic organisms, are highly understudied, despite their unusual biophysical properties. We present here an engineered chimeric PTP (ShufPTP) generated by shuffling the amino acid sequence of five extant hyperthermophilic archaeal PTPs. Despite ShufPTP’s high sequence similarity to its natural counterparts, it presents a suite of unique properties, including high flexibility of the phosphate binding P-loop, facile oxidation of the active-site cysteine, mechanistic promiscuity, and, most notably, hyperthermostability, with a denaturation temperature likely >130 °C (>8 °C higher than the highest recorded growth temperature of any archaeal strain). Our combined structural, biochemical, biophysical, and computational analysis provides insight both into how small steps in evolutionary space can radically modulate the biophysical properties of an enzyme and showcases the tremendous potential of archaeal enzymes for biotechnology, to generate novel enzymes capable of operating under extreme conditions.</p>}},
author = {{Yehorova, Dariia and Alansson, Nikolas and Shen, Ruidan and Denson, Joshua M. and Robinson, Michael and Risso, Valeria A. and Ramirez Molina, Nuria and Loria, J. Patrick and Gaucher, Eric A. and Sanchez-Ruiz, Jose M. and Hengge, Alvan C. and Johnson, Sean J. and Kamerlin, Shina C.L.}},
issn = {{2691-3704}},
keywords = {{archaea; catalytic backups; protein conformational dynamics; protein tyrosine phosphatase; sequence shuffling}},
language = {{eng}},
number = {{1}},
pages = {{59--81}},
publisher = {{The American Chemical Society (ACS)}},
series = {{JACS Au}},
title = {{Conformational Dynamics and Catalytic Backups in a Hyper-thermostable Engineered Archaeal Protein Tyrosine Phosphatase}},
url = {{http://dx.doi.org/10.1021/jacsau.5c00756}},
doi = {{10.1021/jacsau.5c00756}},
volume = {{6}},
year = {{2026}},
}