PDF《Lithium》

40 years ago by Guroff [1]. The cDNA for this protein was isolated in the middle of the 1980s [2], and based on the homology of its sequences there are now

15 *Address correspondence to this author at the Centros de Investigación Biomédica en Red de Enfermedades Neurodegenerativas (CIBERNED), Institut de Biomedicina (IBUB), Unitat de Farmacologia i Farmacognòsia, Facultat de Farmàcia, Universitat de Barcelona, Avda. Diagonal, 643,

08028 Barcelona, Spain;

E-mail: [email protected] different isoforms within the human genome (all the possible box isoforms). Most of them have been identified only as mRNA, and are thought to be tissue specific [3]. Only two small regulatory subunits and 1, 2, 3, 5,

10 catalytic subunit isoforms have been identified in the brain. The best characterized and most abundant calpains in the brain are -and m-calpain. Both proteases exist as heterodimers, each with a definite catalytic subunit of

80 kDa that is encoded by the gene CAPN1 of human chromosome

11 or by the gene CAPN2 of human chromosome

1 respectively, and a small regulatory subunit common to both isoforms. The catalytic subunits share 50% ho- mology and through an analysis of their amino acid sequences four different functional domains can be identified [2] (Fig. 1). Calpain isoform 2, the catalytic subunit of m-calpain and the first to be iso- lated by crystallization [4], contains an anchor domain I at the N- terminal end which suffers a process of autolytic cleavage, probably through internal molecular machinery. The catalytic region in which calpain interacts with its substrates and which contains the protease activity is in domain II. The role of domain III within this calpain is virtually unknown, although there are references in con- nection with its participation in the union to calcium and phosphol- ipids [5]. Domain IV is the calcium-binding domain and contains five EF-hand motifs (reviewed by Bevers et al. 2008). In the ab- sence of calcium, the set formed by the catalytic cysteine, histidine and asparagine residues in domain II are separated from each and other, indicating that a conformational change is required prior to activation [6]. The biggest structural difference between isoforms

1 and 2, - and m- calpain respectively, is that in isoform 1, the separation between the three catalytic residues is smaller and the calcium- binding domain IV is more flexible. This feature could explain the main biochemical difference between - and m-calpain, namely the concentration of calcium required for activation in vitro. -Calpain requires a concentration in the order of 3C50 M for its activation, while m-calpain requires 0.4C0.8 mM calcium. Both calpains share a common regulator subunit, which is sometimes called calpain 4.

2 Current Drug Metabolism, 2009, Vol. 10, No.

5 Camins et al. Table 1. Diseases Associated with Calpain Alterations Disease/Condition Evidence/Comments Genetic diseases Limb girdle muscular dystrophy type 2A Caused by disruptions in the gene for calpain 3;

disease-related disruptions are linked to loss of calpain

3 proteolytic activity. Type II Diabetes mellitus Mutations in intron

3 of Capn10 gene associated with increased incidence of type

2 diabetes in some populations. Ca2+ homeostasis-linked pathologies Alzheimer'

s disease Amount of m-calpain in the cytosolic but not the membranous fractions and in the neurofibrillary tangles of brain from Alzheimer'

s patients is increased. Ratio of autolyzed to unautolyzed -calpain is threefold higher and amount of calpastatin is lower in AD prefrontal cortex than in normal brain. Cataract formation Ca2+ influx activates m-calpain, the predominant calpain in the lens, and C crystallins are cleaved. The crystallin fragments aggregate to form cataracts. Muscular dystrophies Many muscular dystrophies are caused by loss of a membrane-associated protein, dystrophin, which results in increased Ca2+ concentra- tions in muscle, loss of Ca2+ homeostasis, and inappropriate calpain activity. Myocardial infarcts Ca2+ homeostasis is lost in ischemic areas, triggering inappropriate calpain activity. Protein and mRNA levels of first m-calpain and then -calpain increase after a myocardial infarction Multiple sclerosis (MS) (demyelina- tion) All major myelin proteins, the 68- and 200-kDa neurofilament proteins, myelin basic protein, and myelin-associated glycoprotein, are calpain substrates. Neuronal ischemia (stroke) Intracellular [Ca2+ ] increases in ischemic areas, partly due to overactivation of NMDA receptors. Tissue damage in ischemic areas involves both apoptosis and necrosis and the calpains participate in both processes Traumatic spinal cord (brain) injury Injury results in increased intracellular [Ca2+ ]. Levels of calpain-specific 145-kDa -spectrin fragment increased up to 665% within