Covalent binding of food-derived blue pigment phycocyanobilin to bovine β-lactoglobulin under physiological conditions

Abstract

In this study, we investigated structural aspects of covalent binding of food derived blue pigment phycocyanobilin (PCB) to bovine β-lactoglobulin (BLG), major whey protein, by spectroscopic, electrophoretic, mass spectrometry and computational methods. At physiological pH (7.2), we found that covalent pigment binding via free cysteine residue is slow (k = 0.065 min), of moderate affinity (K = 4 × 10 M), and stereo-selective. Binding also occurs at a broad pH range and under simulated gastrointestinal conditions. Adduct formation rises with pH, and in concentrated urea (k = 0.101 min). The BLG-PCB adduct has slightly altered secondary and tertiary protein structure, and bound PCB has higher fluorescence and more stretched conformation than free chromophore. Combination of steered molecular dynamic for disulfide exchange, non-covalent and covalent docking, favours Cys119 residue in protein calyx as target for covalent BLG-PCB adduct formation. Our results suggest that this adduct can serve as delivery system of bioactive PCB.

C-Phycocyanin-derived Phycocyanobilin as a Potential Nutraceutical Approach for Major Neurodegenerative Disorders and COVID-19- induced Damage to the Nervous System

“…Given the versatile and wide use of BSA as a biological carrier of a diverse range of pharmaceutical and nutraceutical compounds, these results point to a possible BSA-PCB stable formulation [ 36 ]. Additionally, PCB also binds to human serum albumin (HSA) [ 37 ] and to bovine β-lactoglobulin (BLG), the main protein composing the milk's whey of cows; this binding was stable in extreme situations, such as an ample pH range and in experimentally modeled gastrointestinal environment [ 38 ]. Interestingly, it was observed that PCB binds to HSA with an affinity similar to BR [ 39 ].…”

Phycocyanobilin reduces brain injury after endothelin‐1‐ induced focal cerebral ischaemia

“…The phycocyanobilin (PCB) chromophore is a linear tetrapyrrole present in the phycobiliproteins C‐phycocyanin and allophycocyanin from the Spirulina platensis cyanobacteria. The chemical structure of PCB is similar to biliverdin, and both could be substrates of biliverdin reductase, an enzyme that transforms these compounds into phycocyanorubin or bilirubin, respectively .…”

Structure of proteins under pressure: Covalent binding effects of biliverdin on β-lactoglobulin

“…This strongly suggests that BLG-BV adducts are not stable in the conditions used in MS/MS. To our knowledge, this is the first time that BLG-BV adducts are detected on a protein by this technique, whereas in a previous study, we were able to detect only one fragment of PCB chromophore bound to BLG but not the whole molecule 11 .…”

“…A similar result was obtained for phycocyanobilin (PCB), a BV analogue, in a previous study on BLG. 11 After HP treatment, the covalent BLG-BV adduct is mostly preserved at the monomeric state (Fig. 2A), whereas BLG without ligand is observed at higher MW (not shown).…”

“…Bovine β-lactoglobulin (BLG), the major whey protein, exhibits good solubility in a broad pH range, as well as numerous techno-functional properties (such as emulsion stabilization and gel formation), which make it a suitable choice for use as an additive in food industry. 10,11 However, pressure treatment has detrimental effects on emulsion efficiency and stability of BLG due to its oligomerization and aggregation. 12 Additionally, BLG is one of the most common allergens of milk 13 , and the effect of HPP on its allergenicity remains unclear and might strongly depend on the pressure values used.…”