9CPM

Thermus thermophilus HB27 laccase (Tth-Lac) mutant with partial deletion of beta-hairpin sequence

Summary for 9CPM

• Entry DOI: 10.2210/pdb9cpm/pdb
• Descriptor: Laccase, COPPER (II) ION, GLYCEROL, ... (4 entities in total)
• Functional Keywords: extremozyme, high-thermal stability, beta-hairpin motif, multicopper oxidase, trinuclear center (tnc), copper coordination, metal binding protein
• Biological source: Thermus thermophilus HB27
• Total number of polymer chains: 1
• Total formula weight: 48759.67

Authors

Lee, J.,Miranda-Zaragoza, B.,Rodriguez-Almazan, C.,Strynadka, N.C.J. (deposition date: 2024-07-18, release date: 2025-02-05)

Primary citation

Miranda-Zaragoza, B.,Huerta-Miranda, G.A.,Garcia-Garcia, W.I.,Hernandez-Alvarez, E.,Solano-Peralta, A.,Lee, J.,Strynadka, N.,Miranda-Hernandez, M.,Rodriguez-Almazan, C.
Structure-Function Relationship of the beta-Hairpin of Thermus thermophilus HB27 Laccase.
Int J Mol Sci, 26:-, 2025

PubMed Abstract: Thermus thermophilus HB27 laccase (Tth-Lac) is a thermostable enzyme that contains a β-hairpin (Ala292-Gln307) covering the substrate entrance. We analyzed the role of this β-hairpin in the enzymatic activity of Tth-Lac through three β-hairpin mutants: two variants without the β-hairpin (C1Tth-Lac and C2Tth-Lac) and one with a partially modified β-hairpin (P1Tth-Lac). Enzymatic activity was assayed with different substrates with and without copper. C1Tth-Lac showed a higher dependency on copper, increasing its activity by 1600-fold for syringaldazine (SGZ). All mutants presented a higher activity than Tth-Lac with phenolic substrates in the presence of copper. The position of the signal associated with CuT2 also changed, as shown in EPR spectra. Elucidation of the crystal structure of P1Tth-Lac mutant (PDB: 9CPM) showed that the partial deletion of the β-hairpin did not significantly affect the overall tertiary structure compared to the wild-type (PDB: 2xu9) nor the coordination of the four internally bound Cu atoms. Higher B-factors of the residues downstream of the deletion indicate increased flexibility (Q307, G308, P309, S310) that were otherwise more ordered in the Tth-Lac structure. Redox potential experiments on platinum electrodes have shown that all proteins have high redox potential, a finding that could have significant implications in the field of protein research.

PubMed: 39859450
DOI: 10.3390/ijms26020735

Experimental method

X-RAY DIFFRACTION (1.6 Å)