Utility of B-Factors in Protein Science: Interpreting Rigidity, Flexibility, and Internal Motion and Engineering Thermostability

Sun, Zhoutong; Liu, Qian; Qu, Ge; Feng, Yan; Reetz, Manfred T.

doi:10.1021/acs.chemrev.8b00290

Cited by 382 publications

(287 citation statements)

References 410 publications

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“…Conformational flexibility has been previously used by existing pathogenic variant callers as one of the many other features (Ancien, Pucci, Godfroid, & Rooman, 2018;Pejaver et al, 2017). B-factor values of atomic coordinates in crystal structures (Sun, Liu, Qu, Feng, & Reetz, 2019), NMR order parameters (Torchia, 2015) or fluctuation calculated from an alignment of multiple X-ray structures, or NMR-derived ensembles allows straightforward estimates on flexibility. Conformational flexibility estimated through MD has also been used as one of the many features in predicting for effects of a missense mutation (Ponzoni & Bahar, 2018), but not in a direct manner as us.…”

Section: Discussionmentioning

confidence: 99%

Exploring the use of molecular dynamics in assessing protein variants for phenotypic alterations

Garg¹,

Pal²

2019

Human Mutation

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With the advent of rapid sequencing technologies, making sense of all the genomic variations that we see among us has been a major challenge. A plethora of algorithms and methods exist that try to address genome interpretation through genotype–phenotype linkage analysis or evaluating the loss of function/stability mutations in protein. Critical Assessment of Genome Interpretation (CAGI) offers an exceptional platform to blind‐test all such algorithms and methods to assess their true ability. We take advantage of this opportunity to explore the use of molecular dynamics simulation as a tool to assess alteration of phenotype, loss of protein function, interaction, and stability. The results show that coarse‐grained dynamics based protein flexibility analysis on 34 CHEK2 and 1719 CALM1 single mutants perform reasonably well for class‐based predictions for phenotype alteration and two‐thirds of the predicted scores return a correlation coefficient of 0.6 or more. When all‐atom dynamics is used to predict altered stability due to mutations for Frataxin protein (8 cases), the predictions are comparable to the state‐of‐the‐art methods. The competitive performance of our straightforward approach to phenotype interpretation contrasts with heavily trained machine learning approaches, and open new avenues to rationally improve genome interpretation.

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Section: Discussionmentioning

confidence: 99%

Exploring the use of molecular dynamics in assessing protein variants for phenotypic alterations

Garg¹,

Pal²

2019

Human Mutation

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“…One approach to compare the relative rigidity and flexibility of proteins in X-ray structural models is to compare the distribution of normalized B-factors in the structures. 33 The normalized B-factor Z-score of the wild type (R303, PDB code: 3DBA) and mutant (R303 C33A/C102A ) structures were calculated. 34 As each X-ray structure contained two molecules in the asymmetric unit, the normalized B-factors for each chain (Chain A and Chain B) in the noncrystallographic symmetry (NCS) dimer were plotted independently to compare global differences in normalized B factors across molecules in the structures, as well as to look specifically at the flexibility of CDR1, CDR2, and CDR3 (Figure 4a).…”

Section: B-factor Analysis Of R303 and R303 C33a/c102amentioning

confidence: 99%

Role of a noncanonical disulfide bond in the stability, affinity, and flexibility of a VHH specific for the Listeria virulence factor InlB

et al. 2020

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A distinguishing feature of camel (Camelus dromedarius) VHH domains are noncanonical disulfide bonds between CDR1 and CDR3. The disulfide bond may provide an evolutionary advantage, as one of the cysteines in the bond is germline encoded. It has been hypothesized that this additional disulfide bond may play a role in binding affinity by reducing the entropic penalty associated with immobilization of a long CDR3 loop upon antigen binding. To examine the role of a noncanonical disulfide bond on antigen binding and the biophysical properties of a VHH domain, we have used the VHH R303, which binds the Listeria virulence factor InlB as a model. Using site directed mutagenesis, we produced a double mutant of R303 (C33A/C102A) to remove the extra disulfide bond of the VHH R303. Antigen binding was not affected by loss of the disulfide bond, however the mutant VHH displayed reduced thermal stability (Tm = 12°C lower than wild‐type), and a loss of the ability to fold reversibly due to heat induced aggregation. X‐ray structures of the mutant alone and in complex with InlB showed no major changes in the structure. B‐factor analysis of the structures suggested that the loss of the disulfide bond elicited no major change on the flexibility of the CDR loops, and revealed no evidence of loop immobilization upon antigen binding. These results suggest that the noncanonical disulfide bond found in camel VHH may have evolved to stabilize the biophysical properties of the domain, rather than playing a significant role in antigen binding.

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“…Our results are in consistent with the previous works that mutate residues near active center improve activity and thermostability. 57 Therefore, we assumed that 38 covariant residues might modulate both functions, which gives a new sight for the diverse co-evolutionary routes.…”

Section: Scasim Reveals Multiple Functions Of Covariant Residuesmentioning

confidence: 99%

Algorithm‐based coevolution network identification reveals key functional residues of the α/β hydrolase subfamilies

Líu

et al. 2019

FASEB j.

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Covariant residues identified by computational algorithms have provided new insights into enzyme evolutionary routes. However, the reliability and accuracy of routine statistical coupling analysis (SCA) are unable to satisfy the needs of protein engineering because SCA depends only on sequence information. Here, we set up a new SCA algorithm, SCA.SIM, by integrating structure information and MD simulation data. The more reliable covariant residues with high‐quality scores are obtained from sequence alignment weighted by residual movement for eight related subfamilies, belonging to α/β hydrolase family, with Candida antarctica lipase B (CALB). The 38 predicted covariant residues are tested for function by high‐throughput quantitative evaluation in combination with activity and thermostability assays of a mutant library and deep sequencing. Based on the landscapes of both activity and thermostability, most mutants play key roles in catalysis, and some mutants gain 2.4‐ to 6‐fold increase in half‐life at 50°C and 9‐ to 12‐fold improvement in catalytic efficiency. The activity of double mutants for A225F/T103A is higher than those of A225F and T103A which means that SCA.SIM method might be useful for identifying the allosteric coupling. The SCA.SIM algorithm can be used for protein coevolution and enzyme engineering research.

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Utility of B-Factors in Protein Science: Interpreting Rigidity, Flexibility, and Internal Motion and Engineering Thermostability

Cited by 382 publications

References 410 publications

Exploring the use of molecular dynamics in assessing protein variants for phenotypic alterations

Exploring the use of molecular dynamics in assessing protein variants for phenotypic alterations

Role of a noncanonical disulfide bond in the stability, affinity, and flexibility of a VHH specific for the Listeria virulence factor InlB

Algorithm‐based coevolution network identification reveals key functional residues of the α/β hydrolase subfamilies

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