Lattice Interactions in Crystals of Soybean Trypsin Inhibitor (Kunitz) Produced by Inclusion of 1,5-Disulfonylnaphthalene

Abstract

In the course of an earlier investigation into the crystallization of proteins based on the addition of intermolecular ligands, the Kunitz type trypsin inhibitor from soybean (SBTI) was crystallized as a complex with 1,5-disulfonylnaphthalene (ligand library 21D). The two molecules within the asymmetric unit of the monoclinic crystals are related by a near-exact NCS 2-fold axis and have essentially the same conformation as was found for them in previous analyses. The protein dimer is maintained through electrostatic interactions with the sulfonyl groups of four 1,5-disulfonylnaphthalene molecules, which are bound in pairs at the dimer interface and involve symmetrical pairs of arg 30 and arg 47 side chains of each SBTI molecule. The binding of the 1,5-disulfonylnaphthalene molecules at the interface is disordered in that only a coordinated single pair of 1,5-disulfonylnaphthalene molecules can be bound at any one time. There are also molecules of the MES buffer present in the crystal structure that are responsible for additional lattice interactions. This crystal structure provides a remarkable demonstration of how unusual, nonphysiologic, small molecules can create symmetry and promote specific lattice interactions within protein crystals.

Noncompetitive tight‐binding inhibition ofAnticarsia gemmatalistrypsins byAdenanthera pavoninaprotease inhibitor affects larvae survival

“…The noncompetitive mechanism found for the inhibition of ApTI on the trypsin‐like enzymes of A. gemmatalis agrees with other in silico studies carried out with trypsins from other species of the order Lepidoptera (Migliolo et al, 2010; Ramalho et al, 2018), and the reactive (inhibitory) peptide bond was identified in the alpha chain with the participation of the Arg64 residue and amino acid residues around Pro63, Ile65, and Arg66. This bond is at an exactly homologous position to the reactive sites identified in the soybean inhibitor at Arg‐63‐Ile‐64 (McPherson, Daym, & Larson, 2019) in Albizia at Arg‐66‐I1e‐67 (Sharma, Nath, Kumari, & Bhardwaj, 2012) and in Psophocarpus at Arg‐64‐Ser‐65 (Yamamoto, Hara, & Ikenaka, 1983).…”

Digestibility of Proteins in Legumes

“…In legumes, the trypsin inhibitor content ranges from 3 to 84 U/mg, while the chymotrypsin inhibitor content varies from 0 to 17 U/mg [53,54]. The prominent trypsin inhibitors in legumes are the Bowman-Birk inhibitor and Kunitz-type inhibitor (Figure 7) [55][56][57]. Kunitz-type inhibitor (molecular weight 18-24 kDa) and Bowman-Birk inhibitors (molecular weight 7-9 kDa) are both capable of inhibiting trypsin and chymotrypsin enzymes.…”