Tryptophan-tryptophan energy transfer and classification of tryptophan residues in proteins using a therapeutic monoclonal antibody as a model.
Intrinsic tryptophan (Trp) fluorescence is often used to determine conformational changes of proteins. The fluorescence of multi-Trp proteins is generally assumed to be additive. This assumption usually holds well if Trp residues are situated at long distances from each other in the absence of any excited state reactions involving these residues and therefore when energy transfer does not occur. Here, we experimentally demonstrate energy transfer among Trp residues and support it by a Master Equation kinetic model applied to a therapeutic monoclonal antibody (mAb). The mAbs are one of the most studied and important biologics for the pharmaceutical industry, and they contain many Trp residues in close proximity. Understanding mAb fluorescence is critical for interpreting fluorescence data and protein-structure relationships. We propose that Trp residues could be categorized into three types of emitters in the mAbs. Experimentally, we categorize them according to solvent accessibility based on dependence of their fluorescence lifetime on the external quencher concentration and their emission wavelength. Theoretically, we categorize with molecular dynamics simulations according to their solvent accessibility. This method of combinatorial mapping of fluorescence characteristics can be utilized to illuminate structural aspects as well as make comparisons of drug formulations for these pharmaceutical proteins.
Kayser
Veysel
V
Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
Chennamsetty
Naresh
N
Voynov
Vladimir
V
Helk
Bernhard
B
Trout
Bernhardt L
BL
D000911:Antibodies, Monoclonal / Q000737:chemistry*; D018360:Crystallography, X-Ray; D031541:Fluorescence Resonance Energy Transfer; D019791:Guanidine; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D011489:Protein Denaturation; D011506:Proteins / Q000737:chemistry*; D014364:Tryptophan / Q000737:chemistry*
D000911:Antibodies, Monoclonal; D011506:Proteins; D014364:Tryptophan; D019791:Guanidine