User:Kevin Buadlart Dagbay

From Proteopedia

Revision as of 20:48, 19 December 2011

Bold textOne of the CBI Molecules being studied in the University of Massachusetts Amherst Chemistry-Biology Interface Program at UMass Amherst and on display at the Molecular Playground.

University of Massachusetts Amherst Protein Regulation/Structural Biology of Caspase

Molecular Playground banner: Caspase-6, a protease that is implicated in neurodegenerative disease including Alzheimer's and Huntington

Molecular Playground banner: Caspase-6 with Z-VAD-FMK, an inhibitor of caspase-6 activity.

Introduction

Cysteine aspartate proteases (caspases) play several key roles in cellular development, homeostasis, and a wide range of diseases. These proteases are normally expressed in cells as inactive precursor zymogens and get activated during processes such as cellular death pathway known as apoptosis [1]. Caspase-6 is particularly interesting since it has been implicated in neurodegenerative diseases including Alzheimer’s and Huntington’s Disease. Alzheimer’s Disease is the major cause of cognitive and cerebral deterioration in older adults. Caspase-6 has been shown to cut amyloid precursor protein (APP), at position 720 leading to the toxic fragment Jcasp, which is one of the fragments possibly responsible for causing the disease morphology [2]. Specific amino acid sequence (IVLD586G) is recognized by caspase-6 in mice with Huntington’s disease that give rise to the development of the behavioral and neuropathological features of the disease [3] [4]. Mutation of the caspase-6 site in mice model with Alzheimer’s and Huntington’s disease provides protection from the neural dysfunction, suggesting a causal relationship between caspase-6 cleavage and neurodegeneration.

Structure and Regulation

The structure of caspase-6 [5] [6] is similar in overall fold to the six other human caspases for which structures are available, all of which are dimeric when active. The structure of ligand-free caspase-6 differs significantly from all other caspases because two novel extended helices are observed flanking the caspase-6 active site. All caspases share a common active-site cysteine–histidine dyad [7] and derive their name, cysteine aspartate proteases, from the presence of the catalytic cysteine at the active site and from their exquisite specificity for cleaving substrate proteins after aspartate residues [8]. Caspases catalyze cleavage of amide bonds via nucleophilic attack of the cysteine thiolate (Cys163 in caspase-6) at the substrate amide carbonyl. During catalysis, the histidine (His121 in caspase-6) activates the catalytic cysteine and a water molecule. Mutation of either of these residues results in loss of catalytic activity [9]. Caspase-6 is expressed as inactive zymogen capable of dimerization. Caspase-6 has three reported cleavage sites that appear to be cleaved by autoproteolysis and or other caspases: D23 in the prodomain, D179 and D193 in the intersubunit linker [10] [[Image:]](see figure below). The prodomain is released from the caspase dimer after cleavage leaving the active form of enzyme consists of two large and two small subunits. The large subunits contain the active site (WOW) catalytic dyad residues, and the small subunits contain most of the dimer interface and the allosteric site (WOW).

The structure of capase-6 (6) The regulation of caspase-6 activity is not well studied. More research on possible ways to regulate caspase-6 activity will provide additional basic understanding of the mechanism of caspase-6 activation. In cells, caspase-6 can be activated by removal of the part of caspase-6, the prodomain, along with cleavage at specific amino acid residue (aspartate 193). In addition, an alternatively spliced form of caspase-6, the caspase beta (C6β) isoform has been shown to inhibit caspase-6 activity. Morover, reported structures of caspase-6 and its inhibitors (Ac-VEID-CHO [PDB ID: 3OD5] & (Z-VAD-FMK [PDB ID: 3QNW]) shed the some basic dynamics of caspase-6 activation including loop rearrangement near the active site (11-12).

Proteins in iLBP family have very high structural conservation despite having very low sequence identities [11]. Similar to other members of the iLBP family, (PDBID: 1CBI) has two orthogonal five-stranded with a between the first and the second β-strands,showing α+β secondary domains. It contains a very large solvent accessible central cavity that binds (PDBID: 1CBS). This conserved is contained between strand 4 and 5 and has no inter-strand hydrogen bonds but is compensated by the presence of ordered water molecules. The helix-turn-helix motif between the first and second strands acts as a on the ligand binding pocket. Strands 7 and 8 are connected by the which has variable lengths within the family. There are 15 fully residues in CRABP I, seven are found in the helix I and II and 5 are in the β-barrel closure [12].

The RA/CRABP interaction is predominantly hydrophobic, as the ligand forms ten contacts with non-polar side chains and only one salt bridge. The β-barrel contains a poorly accessible hydrophobic ligand-binding cavity. For Chain A, the residues in of CRABP I to RA correspond to PRO39,THR56, LEU120, ARG131, and TYR133. In case of Chain B, even though MET 27 is not one of the residues in chain B, it can be one of the the to RA which is bound to PRO39, THR56, LEU120, ARG131, and TYR133 in chain B of CRABP I. Comparison of the and the structure of the proteins in the iLBP family does not reveal a significant opening large enough to allow ligand entry and release [13]. Entry of RA into the cavity of CRABPs is proposed to occur via a region of the protein comprising determinants from the βC-D loop, the βE-F loop and the N-terminal region of helix II. This region of the protein referred to as the “portal” region of the protein has been extensively studied in other members of the iLBP family, in particular in the fatty acid binding protein, by X-ray crystallography, mutational analysis and multidimensional NMR [14].

See Also

• Cellular retinoic acid-binding protein 1 at Wikipedia

3D structure of Cellular retinoic acid-binding protein

CRABP I

2cbr - hCRABP I – human
1cbr - CRABP I + retinoic acid – mouse

CRABP II

2fs6, 2fs7 - hCRABP II
1blr - hCRABP II – NMR
3fek, 3fel, 3fen, 3fa7, 3fa8, 3fa9, 3i17, 3d95, 3d96, 3d97, 2frs - hCRABP II (mutant)
3f8a, 3f9d, 3fa6, 3cr6, 2g79, 2g7b – hCRABP II (mutant) + retinal analog
3cwk, 2g78 – hCRABP II (mutant) + retinoic acid
2fr3, 2cbs, 3cbs, 1cbq, 1cbs – hCRABP II + retinoic acid
3fep - hCRABP II (mutant) + ligand

References

• [1] Venepally, P. et al. Biochemistry. 35, 9974-9982 (1996) PMID: 8756459 [PubMed - indexed for MEDLINE]
• [2] Vaessen, et al. Differentiation. 40, 99-105 (1989). PMID: 2547683 [PubMed - indexed for MEDLINE]
• [3] Gunasekaran, K, "et al". PROTEINS: Structure, Function, and Bioinformatics. PMID: 14696180
• [4] Marcelino, A, "et al". PROTEINS: Structure, Function, and Bioinformatics. PMID: 16477649 [Pubmed- indexed for MEDLINE]
• [5] Xiao, H. and I. A. Kaltashov,J Am Soc Mass Spectrom. 16(6),869-79 (2005). | doi:10.1016/j.jasms.2005.02.020
• [6] Krishnan, V. et al. Biochemistry 39(31), 9119-9129 (2000)PMID: 10924105 [PubMed - indexed for MEDLINE]
• [7] Sacchettini, J. et al. J Biol Chem. 267(33), 23534-23545 (1992)
• [8] Hodsdon, M.et al. Biochemistry. 36(6), 1450-60(1997)| doi:10.1021/bi961890r
• [9] Sjoelund, V. et al. Biochemistry.46, 13382–13390 (2007) | doi: 10.1021/bi700867c