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. 2024 May 15;25(10):5398.
doi: 10.3390/ijms25105398.

Protopine and Allocryptopine Interactions with Plasma Proteins

Aleksandra Marciniak  1 , Aleksandra Kotynia  1 , Edward Krzyżak  1 , Żaneta Czyżnikowska  1 , Sylwia Zielińska  2 , Weronika Kozłowska  2 , Marcel Białas  3 , Adam Matkowski  4 , Anna Jezierska-Domaradzka  2
Affiliations

Protopine and Allocryptopine Interactions with Plasma Proteins

Aleksandra Marciniak et al. Int J Mol Sci. .

Abstract

A comprehensive study of the interactions of human serum albumin (HSA) and α-1-acid glycoprotein (AAG) with two isoquinoline alkaloids, i.e., allocryptopine (ACP) and protopine (PP), was performed. The UV-Vis spectroscopy, molecular docking, competitive binding assays, and circular dichroism (CD) spectroscopy were used for the investigations. The results showed that ACP and PP form spontaneous and stable complexes with HSA and AAG, with ACP displaying a stronger affinity towards both proteins. Molecular docking studies revealed the preferential binding of ACP and PP to specific sites within HSA, with site 2 (IIIA) being identified as the favored location for both alkaloids. This was supported by competitive binding assays using markers specific to HSA's drug binding sites. Similarly, for AAG, a decrease in fluorescence intensity upon addition of the alkaloids to AAG/quinaldine red (QR) complexes indicated the replacement of the marker by the alkaloids, with ACP showing a greater extent of replacement than PP. CD spectroscopy showed that the proteins' structures remained largely unchanged, suggesting that the formation of complexes did not significantly perturb the overall spatial configuration of these macromolecules. These findings are crucial for advancing the knowledge on the natural product-protein interactions and the future design of isoquinoline alkaloid-based therapeutics.

Keywords: albumin; allocryptopine; orosomucoid; protopine; spectroscopy.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
Protopine and allocryptopine molecular structures.
Figure 2
Figure 2
The changes in UV absorption spectra during the titration of (A) HSA by ACP, (B) HSA by PP, (C) AAG by ACP, and (D) AGG by PP.
Figure 3
Figure 3
The UV absorption data analysis plot: the Benesi–Hildebrand procedure for titration of HSA or AAG by (A) ACP (B) PP and the Scatcher plot for titration of HSA or AAG by (C) ACP and (D) PP.
Figure 4
Figure 4
The backbone atoms’ RMSD plots during 100 ns MD simulation for unliganded HSA and with ACP and PP docked at site 1 and site 2.
Figure 5
Figure 5
Rg plots of free HSA and for complex with ACP and PP docked at site 1 and site 2, during 100 ns MD simulation.
Figure 6
Figure 6
The fluorescence spectra of (A) HSA/dGly complex titrated with the studied compound ACP, (B) HSA/dGly complex titrated with the studied compound PP, (C) HSA/dPhe complex titrated with the studied compound ACP, (D) HSA/dPhe complex titrated with the studied compound PP, (E) AAG/QR complex titrated with the studied compound ACP, and (F) AAG/QR complex titrated with the studied compound PP.
Figure 7
Figure 7
Location of allocryptopine (red) and protopine (blue) at the HSA binding site 2.
Figure 8
Figure 8
The 2D plot of interactions between allocryptopine (left) and protopine (right) with HSA at the binding site 2.
Figure 9
Figure 9
The backbone atoms’ RMSD plots during 100 ns MD simulation for AAG, AAG-ACP, and AAG-PP.
Figure 10
Figure 10
Rg plots during 100 ns MD simulation for AAG, AAG-ACP, and AAG-PP.
Figure 11
Figure 11
Location of allocryptopine (red) and protopine (blue) in the AAG pocket.
Figure 12
Figure 12
The 2D plot of interactions between allocryptopine (left) and protopine (right) with AAG.
Figure 13
Figure 13
The circular dichroism spectra of: (A) HSA with ACP, (B) HAS with PP, (C) AAG with ACP, and (D) AAG with PP.
See this image and copyright information in PMC

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