Knob-Socket Predictions of Alpha-Helical Stability and Structure
Poster Number
17B
Format
Poster Presentation
Faculty Mentor Name
Jerry Tsai
Faculty Mentor Department
Chemistry
Graduate Student Mentor Name
Taylor Rabara
Graduate Student Mentor Department
Chemistry
Additional Mentors
Graduate Mentor:
Melina Huey
m_huey@u.pacifc.edu
Chemistry
Abstract/Artist Statement
The Knob-Socket (KS) Model is a novel method to describe protein packing. In this work, the KS model is applied to predict intra-helical and inter-helical packing from 2° through 4° structures involving a four-residue tetrahedral motif. The model involves a four-residue tetrahedral motif called the Knob-Socket. The motif consists of a one amino acid knob from one 2° structure that can be packed into a three amino acid socket on another 2° structure. From an analysis of structures in the Protein Data Bank (PDB), the propensity of a set of three amino acids forming a socket was found to exist in 3 states: (1) free, without a knob and favoring intra-helical interactions, (2) filled, packed with a knob, favoring inter-helical interactions and (3) non, unpacked and disfavoring alpha-helical structure. From these propensities, a parallel, α-helical protein homodimer designated KSα1.1, was designed to validate the Knob-Socket Model. In previous research, single and double point mutations were introduced into the wild-type protein sequence. These mutations’ effect on helix content and stability correlated with the change in propensity of the six socket hexagon or Rabara surrounding a mutation. In this current research, additional single point mutations were introduced into the stable KSα-T14V/M20L mutant DNA. As an amino acid packing code, the calculated socket propensities allow for the prediction of the change in secondary structure content and stability to the KSα1.1-T14V/M20L protein. The mutant variants were created through site-directed mutagenesis and were analyzed through circular dichroism to further characterize the alpha-helicity of these mutants. The raw data collected from CD were then deconvoluted using DICHROWeb to quantify the alpha-helical content of each mutant. The alpha helical character of these mutants was then compared to the original predictions made from the change in propensities of the Rabara hexagon. An increase in alpha-helicity would indicate a more stable structure, whereas, a decrease indicates a less stable structure. To further characterize the stability of these structures, thermal and chemical denaturation studies were carried out for each mutant protein.
Location
DeRosa University Center Ballroom
Start Date
27-4-2018 10:00 AM
End Date
27-4-2018 12:00 PM
Knob-Socket Predictions of Alpha-Helical Stability and Structure
DeRosa University Center Ballroom
The Knob-Socket (KS) Model is a novel method to describe protein packing. In this work, the KS model is applied to predict intra-helical and inter-helical packing from 2° through 4° structures involving a four-residue tetrahedral motif. The model involves a four-residue tetrahedral motif called the Knob-Socket. The motif consists of a one amino acid knob from one 2° structure that can be packed into a three amino acid socket on another 2° structure. From an analysis of structures in the Protein Data Bank (PDB), the propensity of a set of three amino acids forming a socket was found to exist in 3 states: (1) free, without a knob and favoring intra-helical interactions, (2) filled, packed with a knob, favoring inter-helical interactions and (3) non, unpacked and disfavoring alpha-helical structure. From these propensities, a parallel, α-helical protein homodimer designated KSα1.1, was designed to validate the Knob-Socket Model. In previous research, single and double point mutations were introduced into the wild-type protein sequence. These mutations’ effect on helix content and stability correlated with the change in propensity of the six socket hexagon or Rabara surrounding a mutation. In this current research, additional single point mutations were introduced into the stable KSα-T14V/M20L mutant DNA. As an amino acid packing code, the calculated socket propensities allow for the prediction of the change in secondary structure content and stability to the KSα1.1-T14V/M20L protein. The mutant variants were created through site-directed mutagenesis and were analyzed through circular dichroism to further characterize the alpha-helicity of these mutants. The raw data collected from CD were then deconvoluted using DICHROWeb to quantify the alpha-helical content of each mutant. The alpha helical character of these mutants was then compared to the original predictions made from the change in propensities of the Rabara hexagon. An increase in alpha-helicity would indicate a more stable structure, whereas, a decrease indicates a less stable structure. To further characterize the stability of these structures, thermal and chemical denaturation studies were carried out for each mutant protein.