5D8N: Tomato leucine aminopeptidase mutant - K354E

Citation:
Abstract
Tomato plants express acidic leucine aminopeptidase (LAP-A) in response to various environmental stressors. LAP-A not only functions as a peptidase for diverse peptide substrates, but also displays chaperone activity. A K354E mutation has been shown to abolish the peptidase activity but to enhance the chaperone activity of LAP-A. To better understand this moonlighting function of LAP-A, the crystal structure of the K354E mutant was determined at 2.15 A resolution. The structure reveals that the K354E mutation destabilizes an active-site loop and causes significant rearrangement of active-site residues, leading to loss of the catalytic metal-ion coordination required for the peptidase activity. Although the mutant was crystallized in the same hexameric form as wild-type LAP-A, gel-filtration chromatography revealed an apparent shift from the hexamer to lower-order oligomers for the K354E mutant, showing a mixture of monomers to trimers in solution. In addition, surface-probing assays indicated that the K354E mutant has more accessible hydrophobic areas than wild-type LAP-A. Consistently, computational thermodynamic estimations of the interfaces between LAP-A monomers suggest that increased exposure of hydrophobic surfaces occurs upon hexamer breakdown. These results suggest that the K354E mutation disrupts the active-site loop, which also contributes to the hexameric assembly, and destabilizes the hexamers, resulting in much greater hydrophobic areas accessible for efficient chaperone activity than in the wild-type LAP-A.
PDB ID: 5D8NDownload
MMDB ID: 139121
PDB Deposition Date: 2015/8/17
Updated in MMDB: 2017/11
Experimental Method:
x-ray diffraction
Resolution: 2.15  Å
Source Organism:
Similar Structures:
Biological Unit for 5D8N: hexameric; determined by author and by software (PISA)
Molecular Components in 5D8N
Label Count Molecule
Proteins (6 molecules)
6
Leucine Aminopeptidase 1, Chloroplastic(Gene symbol: LAPA1)
Molecule annotation
Chemicals (46 molecules)
1
16
2
12
3
4
4
14
* Click molecule labels to explore molecular sequence information.

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