Molecular dynamics simulations reveal a new role for a conserved active site asparagine in a ubiquitin-conjugating enzyme

Wilson, R. Hunter; Zamfir, Serban; Sumner, Isaiah

doi:10.1016/j.jmgm.2017.07.006

Cited by 10 publications

(39 citation statements)

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“…The need to reexamine the role of the conserved asparagine is also apparent from the structural data available from NMR and crystal structures of E2~Ub and E2~SUMO conjugates. Contrary to expectations, several of these structures do not show the conserved asparagine forming a hydrogen bond with the thioester carbonyl . (The thioester carbonyl develops the negative charge in the intermediate.)…”

Section: Introductioncontrasting

confidence: 84%

“…There is evidence to support this claim. First, it is part of the HPN loop, a group of residues (histidine, proline, and asparagine) that are highly conserved across the entire E2 family . Second, if the asparagine is mutated, the catalytic rate and efficiency of ubiquitination decrease significantly .…”

Section: Introductionmentioning

confidence: 99%

“…Third, as shown by X‐ray crystallography and computation, it is the only plausible residue capable of acting as an oxyanion hole. However, there have been some recent studies that have reexamined this assumption using crystallography and molecular dynamics (MD) and found evidence that the asparagine hydrogen bonds to a nearby loop in the E2 enzyme, which may affect the loop's dynamics. Mutations to this active site loop are also known to have a negative impact on catalysis and enzyme kinetics, NMR relaxation, and MD experiments have suggested that this loop regulates E2 activity by acting as a gate …”

Section: Introductionmentioning

confidence: 99%

“…In fact, the asparagine side chain is sometimes oriented in the opposite direction. Moreover, studies have shown that the relative position of Ub to the E2 is quite mobile and that the relative position seems to affect the hydrogen bonding partners of the conserved asparagine . Of the possible E2~Ub orientations, the closed or folded back position is thought to be the most active .…”

Section: Introductionmentioning

confidence: 99%

“…In an attempt to understand some of these mechanistic details, we recently used classical MD simulations to examine the role of the conserved asparagine (Asn79) in the E2 enzyme, Ubc13 . We ran simulations of Ub conjugated with WT and mutated (N79A and N79D) Ubc13.…”

Section: Introductionmentioning

confidence: 99%

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A conserved asparagine in a ubiquitin‐conjugating enzyme positions the substrate for nucleophilic attack

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The mechanism used by the ubiquitin-conjugating enzyme, Ubc13, to catalyze ubiquitination is probed with three computational techniques: Born-Oppenheimer molecular dynamics, single point quantum mechanics/molecular mechanics energies, and classical molecular dynamics. These simulations support a long-held hypothesis and show that Ubc13-catalyzed ubiquitination uses a stepwise, nucleophilic attack mechanism. Furthermore, they show that the first step-the formation of a tetrahedral, zwitterionic intermediate-is rate limiting. However, these simulations contradict another popular hypothesis that supposes that the negative charge on the intermediate is stabilized by a highly conserved asparagine (Asn79 in Ubc13). Instead, calculated reaction profiles of the N79A mutant illustrate how charge stabilization actually increases the barrier to product formation. Finally, an alternate role for Asn79 is suggested by simulations of wild-type, N79A, N79D, and H77A Ubc13: it stabilizes the motion of the electrophile prior to the reaction, positioning it for nucleophilic attack.

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Section: Introductioncontrasting

confidence: 84%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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A conserved asparagine in a ubiquitin‐conjugating enzyme positions the substrate for nucleophilic attack

et al. 2019

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Twenty years of exceptional success: The molecular education and research consortium in undergraduate computational chemistry (MERCURY)

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2020

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The molecular education and research consortium in undergraduate computational chemistry (MERCURY) consortium, established in 2000, has contributed greatly to the scientific development of faculty and undergraduates. The MERCURY faculty peer-reviewed publication rate from 2001 to 2019 of 1.7 papers/faculty/year is 3.4 times the rate of the physical science faculty at primarily undergraduate institutions. We have worked with over 1000 students on research projects since 2001, and 75% of our undergraduate research students have been under-represented in chemistry, either female or students of color. Approximately half of our alumni attend graduate school for the purpose of obtaining advanced degrees in STEM fields, and two-thirds are female and/or students of color. We have had more than 1600 attendees at 18 MERCURY conferences, including 111 invited speakers, 61 of whom have been female and/or faculty of color. In this paper, the research accomplishments, transformational outcomes, and scientific productivity of the MERCURY faculty are highlighted.

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A comprehensive in silico analysis of sortase superfamily

Malik

Kim

2019

J Microbiol.

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Molecular dynamics simulations reveal a new role for a conserved active site asparagine in a ubiquitin-conjugating enzyme

Cited by 10 publications

References 50 publications

A conserved asparagine in a ubiquitin‐conjugating enzyme positions the substrate for nucleophilic attack

A conserved asparagine in a ubiquitin‐conjugating enzyme positions the substrate for nucleophilic attack

Twenty years of exceptional success: The molecular education and research consortium in undergraduate computational chemistry (MERCURY)

A comprehensive in silico analysis of sortase superfamily

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