What question did this study set out to answer?

To determine whether electrostatic interactions involving loop 1 in human beta-cardiac myosin regulate the kinetics of ATP binding, actomyosin dissociation, and ADP release.

October 9, 2021Open Access

Loop 1 of human cardiac myosin regulates the rate of ATP induced actomyosin dissociation but not the rate of ADP release

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Why the study?

It was hypothesized that electrostatic interactions between loop 1 and the myosin head backbone regulate ATP binding to and ADP dissociation from actomyosin.

Key Points

To determine whether electrostatic interactions involving loop 1 in human beta-cardiac myosin regulate the kinetics of ATP binding, actomyosin dissociation, and ADP release.
Engineered a D208Q:K450L double mutation in human beta-cardiac myosin head constructs (residues 1–843) to disrupt the loop 1-to-backbone salt bridge.
Expressed wild-type and mutant myosin head proteins in differentiated mouse C2C12 cells.
Measured the kinetics of ATP-induced actomyosin dissociation and ADP release using transient kinetics spectrophotometry.
Mutant actomyosin exhibited a faster rate of ATP-induced dissociation compared to the wild type, demonstrating that loop 1 flexibility modulates ATP binding kinetics.
The rate of ADP release from actomyosin remained unchanged between the mutant and wild type, indicating loop 1 does not regulate the ADP dissociation step.

Structured PICO

Population

Recombinant human cardiac myosin (wild type beta isoform and D208Q:K450L mutant, 1-843 amino acid residues) expressed in C2C12 mouse myoblast cells, interacting with actin from rabbit skeletal tissue.

Intervention

Double mutation D208Q:K450L in the beta isoform of human cardiac myosin to remove the salt bridge D208:K450.

Comparator

Wild type beta isoform of human cardiac myosin.

Outcome

Kinetics of ATP-induced actomyosin dissociation and ADP release measured using transient kinetics spectrophotometry.surrogate

The flexibility of loop 1 in human cardiac myosin regulates the rate of ATP-induced actomyosin dissociation but does not affect the rate of ADP release.

Limitations

Discrepancy of the fits for the WT actomyosin and the mutant
High standard deviation of the rate for the mutant

Discussion

No takes yet. Share an insight, caveat, or question.

Implication

Tests myosin loop 1 salt bridge in nucleotide binding; extends MD simulations but leaves open effects on cardiac function.

Cite This Study

Gargey et al. (2021) studied this question.

synapsesocial.com/papers/6a7372e720644e164b4e5b7e https://doi.org/10.1101/2021.10.08.463735

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Loop 1 of human cardiac myosin regulates the rate of ATP induced actomyosin dissociation but not the rate of ADP release

Key Points

Structured PICO

Limitations

Discussion

Implication

Cite This Study

Loop 1 of human cardiac myosin regulates the rate of ATP induced actomyosin dissociation but not the rate of ADP release

Key Points

Structured PICO

Limitations

Discussion

Implication

Cite This Study

Also Consider

Also Consider