Active subsite properties, subsite residues and targeting to lysosomes or midgut lumen of cathepsins L from the beetle Tenebrio molitor

Active subsite properties, subsite residues and targeting to lysosomes or midgut lumen of cathepsins L from the beetle Tenebrio molitor

Author Damasceno, Ticiane F. Google Scholar
Dias, Renata O. Google Scholar
de Oliveira, Juliana R. Autor UNIFESP Google Scholar
Salinas, Roberto K. Google Scholar
Juliano, Maria A. Autor UNIFESP Google Scholar
Ferreira, Clelia Google Scholar
Terra, Walter R. Google Scholar
Abstract Cathepsins L are the major digestive peptidases in the beetle Tenebrio molitor. Two digestive cathepsins L (TmCAL2 and TmCAL3) from it had their 3D structures solved. The aim of this paper was to study in details TmCAL3 specificity and properties and relate them to its 3D structure. Recombinant TmCAL3 was assayed with 64 oligopeptides with different amino acid replacements in positions P2, P1, P1' and P2'. Results showed that TmCAL3 S2 specificity differs from the human enzyme and that its specificities also explain why on autoactivation two propeptide residues remain in the enzyme. Data on free energy of binding and of activation showed that S1 and S2' are mainly involved in substrate binding, S1' acts in substrate binding and catalysis, whereas 52 is implied mainly in catalysis. Enzyme subsite residues were identified by docking with the same oligopeptide used for kinetics. The subsite hydrophobicities were calculated from the efficiency of hydrolysis of different amino acid replacements in the peptide and from docking data. The results were closer for S1 and S2' than for SI and S2, indicating that the residue subsites that were more involved in transition state binding are different from those binding the substrate seen in docking. Besides TmCAL1-3, there are nine other cathepsins L, most of them more expressed at midgut. They are supposed to be directed to lysosomes by a Drosophila-like Lerp receptor and/or motifs in their prodomains. The mannose 6-phosphate lysosomal sorting machinery is absent from T. molitor transcriptome. Cathepsin L direction to midgut contents seems to depend on overexpression. 2017 Elsevier Ltd. All rights reserved.
Keywords Cathepsin L-like peptidase
Substrate specificity
Subsite role
Subsite hydrophobicity
Phosphotransferase
Cathepsin L activation
Tenebrio molitor
xmlui.dri2xhtml.METS-1.0.item-coverage Oxford
Language English
Sponsor Brazilian research agency FAPESP
Brazilian research agency CNPq
CNPq
FAPESP
Grant number FAPESP: 2011/51685-8
CNPq: 132586/2012-6
CNPq: 142090/2014-0
CNPq: 467478/2014-7
FAPESP 2015/19541-7
FAPESP: 2016/07490-1
Date 2017
Published in Insect Biochemistry And Molecular Biology. Oxford, v. 89, p. 17-30, 2017.
ISSN 0965-1748 (Sherpa/Romeo, impact factor)
Publisher Pergamon-Elsevier Science Ltd
Extent 17-30
Origin http://dx.doi.org/10.1016/j.ibmb.2017.08.004
Access rights Closed access
Type Article
Web of Science ID WOS:000413177300003
URI https://repositorio.unifesp.br/handle/11600/57284

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