PDBsum entry 4cvw

Hydrolase/hydrolase inhibitor

PDB id: 4cvw

Contents

Protein chains

874 a.a.

101 a.a.

Metals

_CA ×4

Waters ×98

PDB id:

4cvw

Name:

Hydrolase/hydrolase inhibitor

Title:

Structure of the barley limit dextrinase-limit dextrinase inhibitor complex

Structure:

Limit dextrinase. Chain: a, b. Fragment: residues 22-904. Engineered: yes. Mutation: yes. Limit dextrinase inhibitor. Chain: c, d. Fragment: residues 25-138. Engineered: yes

Source:

Hordeum vulgare. Barley. Organism_taxid: 4513. Expressed in: komagataella pastoris. Expression_system_taxid: 644223.

Resolution:

2.67Å R-factor: 0.257 R-free: 0.293

Authors:

M.S.Moeller,M.B.Vester-Christensen,J.M.Jensen,M.Abou Hachem, A.Henriksen,B.Svensson

Key ref:

M.S.Møller et al. (2015). Crystal structure of barley limit dextrinase-limit dextrinase inhibitor (LD-LDI) complex reveals insights into mechanism and diversity of cereal type inhibitors. J Biol Chem, 290, 12614-12629. PubMed id: 25792743 DOI: 10.1074/jbc.M115.642777

Date:

31-Mar-14 Release date: 01-Apr-15

Protein chains

O48541  (O48541_HORVV) -  Limit dextrinase from Hordeum vulgare subsp. vulgare

Seq: 904 a.a.

Struc: 874 a.a.*

Protein chains

Q2V8X0  (Q2V8X0_HORVU) -  Limit dextrinase inhibitor from Hordeum vulgare

Seq: 147 a.a.

Struc: 101 a.a.

* PDB and UniProt seqs differ at 4 residue positions (black crosses)

Enzyme reactions

• Enzyme class: Chains A, B: E.C.3.2.1.41 - pullulanase.
• Reaction: Hydrolysis of (1->6)-alpha-D-glucosidic linkages in pullulan and in amylopectin and glycogen, and the alpha- and beta-limit dextrins of amylopectin and glycogen.

DOI no: 10.1074/jbc.M115.642777

J Biol Chem 290:12614-12629 (2015)

PubMed id: 25792743

Crystal structure of barley limit dextrinase-limit dextrinase inhibitor (LD-LDI) complex reveals insights into mechanism and diversity of cereal type inhibitors.

M.S.Møller, M.B.Vester-Christensen, J.M.Jensen, M.A.Hachem, A.Henriksen, B.Svensson.

ABSTRACT

Molecular details underlying regulation of starch mobilization in cereal seed endosperm remain unknown despite the paramount role of this process in plant growth. The structure of the complex between the starch debranching enzyme barley limit dextrinase (LD), hydrolyzing α-1,6-glucosidic linkages, and its endogenous inhibitor (LDI) was solved at 2.7 Å. The structure reveals an entirely new and unexpected binding mode of LDI as compared with previously solved complex structures of related cereal type family inhibitors (CTIs) bound to glycoside hydrolases but is structurally analogous to binding of dual specificity CTIs to proteases. Site-directed mutagenesis establishes that a hydrophobic cluster flanked by ionic interactions in the protein-protein interface is vital for the picomolar affinity of LDI to LD as assessed by analysis of binding by using surface plasmon resonance and also supported by LDI inhibition of the enzyme activity. A phylogenetic analysis identified four LDI-like proteins in cereals among the 45 sequences from monocot databases that could be classified as unique CTI sequences. The unprecedented binding mechanism shown here for LDI has likely evolved in cereals from a need for effective inhibition of debranching enzymes having characteristic open active site architecture. The findings give a mechanistic rationale for the potency of LD activity regulation and provide a molecular understanding of the debranching events associated with optimal starch mobilization and utilization during germination. This study unveils a hitherto not recognized structural basis for the features endowing diversity to CTIs.