Limits...
Determining the role of missense mutations in the POU domain of HNF1A that reduce the DNA-binding affinity: A computational approach

View Article: PubMed Central - PubMed

ABSTRACT

Maturity-onset diabetes of the young type 3 (MODY3) is a non-ketotic form of diabetes associated with poor insulin secretion. Over the past years, several studies have reported the association of missense mutations in the Hepatocyte Nuclear Factor 1 Alpha (HNF1A) with MODY3. Missense mutations in the POU homeodomain (POUH) of HNF1A hinder binding to the DNA, thereby leading to a dysfunctional protein. Missense mutations of the HNF1A were retrieved from public databases and subjected to a three-step computational mutational analysis to identify the underlying mechanism. First, the pathogenicity and stability of the mutations were analyzed to determine whether they alter protein structure and function. Second, the sequence conservation and DNA-binding sites of the mutant positions were assessed; as HNF1A protein is a transcription factor. Finally, the biochemical properties of the biological system were validated using molecular dynamic simulations in Gromacs 4.6.3 package. Two arginine residues (131 and 203) in the HNF1A protein are highly conserved residues and contribute to the function of the protein. Furthermore, the R131W, R131Q, and R203C mutations were predicted to be highly deleterious by in silico tools and showed lower binding affinity with DNA when compared to the native protein using the molecular docking analysis. Triplicate runs of molecular dynamic (MD) simulations (50ns) revealed smaller changes in patterns of deviation, fluctuation, and compactness, in complexes containing the R131Q and R131W mutations, compared to complexes containing the R203C mutant complex. We observed reduction in the number of intermolecular hydrogen bonds, compactness, and electrostatic potential, as well as the loss of salt bridges, in the R203C mutant complex. Substitution of arginine with cysteine at position 203 decreases the affinity of the protein for DNA, thereby destabilizing the protein. Based on our current findings, the MD approach is an important tool for elucidating the impact and affinity of mutations in DNA-protein interactions and understanding their function.

No MeSH data available.


The radius of gyration analysis of native and mutant complexes.Color scheme: native (black), R131W (red), R131Q (green), and R203C (blue). The native complex exhibited the least Rg value, whereas the R203C mutant complex exhibited the highest Rg value. R131W and R131Q mutant complexes showed Rg values between the native and R203C mutant complex values.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC5391926&req=5

pone.0174953.g006: The radius of gyration analysis of native and mutant complexes.Color scheme: native (black), R131W (red), R131Q (green), and R203C (blue). The native complex exhibited the least Rg value, whereas the R203C mutant complex exhibited the highest Rg value. R131W and R131Q mutant complexes showed Rg values between the native and R203C mutant complex values.

Mentions: The radius of gyration (Rg) was utilized to measure and understand the compactness of the protein complexes (Fig 6). The lowest Rg value of ~0.25nm was observed for the native complex. Among the three mutant complexes, the least compactness was observed for the R203C mutant complex, with an Rg value of~2.35nm, followed by the R131W and R131Q mutant complexes with Rg value ~0.3nm, which exhibited similar deviation patterns and convergence. The increased Rg value may also explain the loss of intermolecular hydrogen bonds as shown in Fig 5. The loss of compactness results from the reduction in the interaction pattern between the protein and DNA. Contact map analysis was conducted using g_mdmat to investigate this phenomenon. In the contact map analysis, a loss of contact in the anti-parallel beta sheet region was observed in all the mutant complexes, confirming the increased destabilizing effect upon mutation compared to the native (S5A–S5D Fig). Based on the results of the RMSD analysis, the intermolecular hydrogen bond formation, Rg values and contact map analysis, the R203C mutant complex exhibits the fewest interactions between the protein and DNA.


Determining the role of missense mutations in the POU domain of HNF1A that reduce the DNA-binding affinity: A computational approach
The radius of gyration analysis of native and mutant complexes.Color scheme: native (black), R131W (red), R131Q (green), and R203C (blue). The native complex exhibited the least Rg value, whereas the R203C mutant complex exhibited the highest Rg value. R131W and R131Q mutant complexes showed Rg values between the native and R203C mutant complex values.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5391926&req=5

pone.0174953.g006: The radius of gyration analysis of native and mutant complexes.Color scheme: native (black), R131W (red), R131Q (green), and R203C (blue). The native complex exhibited the least Rg value, whereas the R203C mutant complex exhibited the highest Rg value. R131W and R131Q mutant complexes showed Rg values between the native and R203C mutant complex values.
Mentions: The radius of gyration (Rg) was utilized to measure and understand the compactness of the protein complexes (Fig 6). The lowest Rg value of ~0.25nm was observed for the native complex. Among the three mutant complexes, the least compactness was observed for the R203C mutant complex, with an Rg value of~2.35nm, followed by the R131W and R131Q mutant complexes with Rg value ~0.3nm, which exhibited similar deviation patterns and convergence. The increased Rg value may also explain the loss of intermolecular hydrogen bonds as shown in Fig 5. The loss of compactness results from the reduction in the interaction pattern between the protein and DNA. Contact map analysis was conducted using g_mdmat to investigate this phenomenon. In the contact map analysis, a loss of contact in the anti-parallel beta sheet region was observed in all the mutant complexes, confirming the increased destabilizing effect upon mutation compared to the native (S5A–S5D Fig). Based on the results of the RMSD analysis, the intermolecular hydrogen bond formation, Rg values and contact map analysis, the R203C mutant complex exhibits the fewest interactions between the protein and DNA.

View Article: PubMed Central - PubMed

ABSTRACT

Maturity-onset diabetes of the young type 3 (MODY3) is a non-ketotic form of diabetes associated with poor insulin secretion. Over the past years, several studies have reported the association of missense mutations in the Hepatocyte Nuclear Factor 1 Alpha (HNF1A) with MODY3. Missense mutations in the POU homeodomain (POUH) of HNF1A hinder binding to the DNA, thereby leading to a dysfunctional protein. Missense mutations of the HNF1A were retrieved from public databases and subjected to a three-step computational mutational analysis to identify the underlying mechanism. First, the pathogenicity and stability of the mutations were analyzed to determine whether they alter protein structure and function. Second, the sequence conservation and DNA-binding sites of the mutant positions were assessed; as HNF1A protein is a transcription factor. Finally, the biochemical properties of the biological system were validated using molecular dynamic simulations in Gromacs 4.6.3 package. Two arginine residues (131 and 203) in the HNF1A protein are highly conserved residues and contribute to the function of the protein. Furthermore, the R131W, R131Q, and R203C mutations were predicted to be highly deleterious by in silico tools and showed lower binding affinity with DNA when compared to the native protein using the molecular docking analysis. Triplicate runs of molecular dynamic (MD) simulations (50ns) revealed smaller changes in patterns of deviation, fluctuation, and compactness, in complexes containing the R131Q and R131W mutations, compared to complexes containing the R203C mutant complex. We observed reduction in the number of intermolecular hydrogen bonds, compactness, and electrostatic potential, as well as the loss of salt bridges, in the R203C mutant complex. Substitution of arginine with cysteine at position 203 decreases the affinity of the protein for DNA, thereby destabilizing the protein. Based on our current findings, the MD approach is an important tool for elucidating the impact and affinity of mutations in DNA-protein interactions and understanding their function.

No MeSH data available.