Year: 2025
Type: article
Abstract: We here present a novel and sensitive methodology for determining the melting point (MP) of Bovine Serum Albumin (BSA) from micromolar to picomolar concentration levels under label free conditions. At 1 pM we could model the melting with a sharp gaussian. However, from the transient state observed during the melting process by using a simple exponential decay model we determined a time constant of 67 seconds. We applied this methodology by studying a 3.3 pM sample of a botulinum toxin A (BoNT-A) (stabilized with 2.8 nanomolar denatured Human Serum Albumin (HSA)). We were able to determine the Tm of BoNT-A in the presence of the approximately 1000-fold more concentrated HSA. Entry for the Table of Contents Protein label-free melting point (MP) determination at ultralow concentrations is a huge problem which concern to the biopharmaceutical industry. Here, we present a novel method to determine the MP of bovine serum albumin (BSA) from 1μM to 1pM under label-free conditions. The benefits of this study match the purposes of stability studies in formulations, in which the protein active component is successful at very low concentrations, such as botulinum toxin A (BoNT-A). We used BOCOUTRE, a commercially available pharmaceutical product based on BoNT-A, and Human Serum Albumin (HSA) as a stabilizer. Our method can detect the MP of the stabilizer protein, even if its concentration is markedly different from that of the active component protein (1000-fold) in the case of BOCOUTRE.
Source: Biomolecules
DOI: 10.3390/biom15111579
Institution: Aalborg University Hospital
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $3,258
OpenAlex ID: W4416092511