title: Molecular Investigation of the Mechanism of Non-Enzymatic Hydrolysis of Proteins and the Predictive Algorithm for Susceptibility.
abstract: A number of potential degradation routes can limit the shelf life of a biotherapeutic. While these are experimentally measurable, the tests to do so require a substantial investment in both time and material, resources rarely available early in the drug development process. To address the potential degradation route of non-enzymatic hydrolysis, we performed a molecular modeling analysis, together with an experimental study, to gain detailed insight into the reaction. On the basis of the mechanism, an algorithm for predicting the likely cleavage sites of a protein has been created. This algorithm measures four key properties during a molecular dynamics simulation, which relate to the key steps of the hydrolysis mechanism, in particular the rate-determining step (which can vary depending on the local environment). The first two properties include the secondary structure and the surface exposure of the amide bond, both of which help detect if the addition of the proton to the amide bond is possible. The second two properties relate to whether the side chain can cyclize and form a furane ring. These two properties are the orientation of the side chain relative to the amide bond and the number of hydrogen bonds between the side chain and the surrounding protein. Overall, the algorithm performs well at identifying reactive versus nonreactive bonds. The algorithm correctly classifies nearly 90% of all amide bonds following an aspartic or glutamic acid residue as reactive or nonreactive.
authors.affiliation: Department of Chemical Engineering, Massachusetts Institute of Technology , Cambridge, Massachusetts 02142, United States.
authors.forename: Timothy M
authors.initials: TM
authors.lastname: Lauer
authors.affiliation: Department of Chemical Engineering, Massachusetts Institute of Technology , Cambridge, Massachusetts 02142, United States.
authors.forename: Geoffrey P F
authors.initials: GP
authors.lastname: Wood
authors.affiliation: Department of Formulation Sciences, MedImmune LLC , Gaithersburg, Maryland 20878, United States.
authors.forename: David
authors.initials: D
authors.lastname: Farkas
authors.affiliation: Department of Formulation Sciences, MedImmune LLC , Gaithersburg, Maryland 20878, United States.
authors.forename: Hasige A
authors.initials: HA
authors.lastname: Sathish
authors.affiliation: Department of Formulation Sciences, MedImmune LLC , Gaithersburg, Maryland 20878, United States.
authors.forename: Hardeep S
authors.initials: HS
authors.lastname: Samra
authors.affiliation: Department of Chemical Engineering, Massachusetts Institute of Technology , Cambridge, Massachusetts 02142, United States.
authors.forename: Bernhardt L
authors.initials: BL
authors.lastname: Trout
mesh_terms: D000465:Algorithms; D001224:Aspartic Acid / Q000737:chemistry*; D018698:Glutamic Acid / Q000737:chemistry*; D006801:Humans; D006860:Hydrogen Bonding; D006868:Hydrolysis; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry*; D013816:Thermodynamics
chemical_list: D011506:Proteins; D001224:Aspartic Acid; D018698:Glutamic Acid