|BS||Biology/Chemistry||University of Mysore||India|
|MS||Biochemistry||University of Mysore||India|
|PhD||Biochemistry||University of Mysore||India|
Structure-function of crystallins, role of ocular proteases and molecular basis for cataract development.
The lens of the eye is an excellent model for studying the effects of aging. The lens is primarily composed of long-lived highly stable proteins called crystallins. The crystallins account for approximately 95% of lens proteins. There are three types of lens crystallins: alpha, beta and gamma. The normally transparent lens often gradually becomes cloudy with aging, leading to cataract formation. Cataract is a major cause of blindness worldwide. By age 80, more than half of Americans either have a cataract or have had cataract surgery. Cataract primarily develops as a result of extensive modification, aggregation and precipitation of the lens crystallins. In our studies of the molecular mechanisms of cataract formation, we are investigating the role of cellular enzymes called proteases and crystallin-derived peptides in cataractogenesis and the structure and function of the major lens crystallin, alpha-crystallin.
Every cataract lens analyzed thus far in laboratories across the world has exhibited evidence of proteolysis (the degradation of crystallin proteins). However, the specific proteases responsible for the proteolysis of lens crystallins are yet to be characterized. Using specific peptide substrates that mimic the in vivo cleavage sites in crystallin, we have demonstrated in lens extracts the presence of proteases that may be responsible for the breakdown of lens proteins. These studies will enable us to develop strategies to control the crystallin degradation in vivo, which may eventually lead to the development of interventions for age-related cataract formation.
Alpha crystallin is an ologomeric protein with a molecular weight of about 800 kDa. It is formed by the subunits A and B, each with a molecular weight of 20 kDa. These subunits have high sequence homology to small heat shock proteins. They display chaperone-like properties. The chaperone-like properties of alpha-crystallin likely plays significant role in preventing the protein aggregation and light scattering that are associated with clouding of the lens, thereby helping to maintain lens transparency. Studies in our laboratory are directed toward the understanding the chaperone-like activity of both A and B subunits of alpha-crystallin. We have also chemically synthesized two peptides, called mini-alpha A-crystallin and mini-alpha B-crystallin, and have shown that these peptides possess the anti-aggregation properties of molecular chaperones. The availability of mini-alpha A and mini-alpha B gives us an opportunity to investigate the structural requirements of the chaperone site as well as the conformational specificity (structure or shape) of target proteins during chaperone action. We are also using peptide chaperones to stabilize mutant crystallins and denaturing proteins which otherwise aggregate and precipitate. These studies may lead to the development of peptide chaperones as therapeutic agents.
While the primary sequence of alpha-crystallin subunits has been known for many years, the tertiary and quaternary organization of alpha-crystallin remains to be elucidated fully. We are studying the structural organization of wild-type and mutant alpha-crystallin subunits using site-directed cysteine mutagenesis and novel isotope-tagged crosslinkers. These studies will help us learn how cataracts develop and how they can be prevented.
Notable Honors and Service
- Fellow, Association for Research in Vision and Opthalmology
- Fellow, American Association for the Advancement of Science
- Academic Editor, PLoS ONE, 2012-
- Member, National Eye Advisory Council, 2011-2014
- Program Committee Chair for Lens Section, Annual Meeting of Association for Research in Vision and Ophthalmology, 2007
- Lew R. Wasserman Merit Award from Research to Prevent Blindness, 2002
- Cataract Research Award from National Foundation for Eye Research, 2000
- Robert E. McCormick Scholar Award from Research to Prevent Blindness, 1999
Raju M, Santhoshkumar P, Sharma KK. (2018). Cell-penetrating Chaperone Peptide Prevents Protein Aggregation And Protects Against Cell Apoptosis. Adv Biosyst. 2(1). doi: 10.1002/adbi.201700095. [PubMed]
Phadte AS, Santhoshkumar P, Sharma KK. (2018). Characterization of an N-terminal mutant of αA-crystallin αA-R21Q associated with congenital cataract. Exp Eye Res. 174:185-195. doi: 10.1016/j.exer.2018.05.016. [PubMed]
Phadte AS, Santhoshkumar P, Sharma KK. (2018). αA-crystallin-derived minichaperone stabilizes αAG98R-crystallin by affecting its zeta potential. Mol Vis. 24:297-304. eCollection 2018. [PubMed]
Raju M, Santhoshkumar P, Sharma KK. (2017). Lens Endogenous Peptide αA66-80 Generates Hydrogen Peroxide and Induces Cell Apoptosis. Aging Dis. 8(1):57-70. doi: 10.14336/AD.2016.0805. eCollection 2017 Feb. [PubMed]
Santhoshkumar P, Karmakar S, Sharma KK. (2016). Structural and functional consequences of chaperone site deletion in αA-crystallin. Biochim Biophys Acta. 1864(11):1529-38. doi: 10.1016/j.bbapap.2016.08.006. [PubMed]
Sharma KK. (2016). Crystallin biochemistry in health and disease. Biochim Biophys Acta. 1860(1 Pt B):147-8. doi: 10.1016/j.bbagen.2015.10.018. [PubMed]
Raju M, Mooney BP, Thakkar KM, Giblin FJ, Schey KL, Sharma KK. (2015). Role of αA-crystallin-derived αA66-80 peptide in guinea pig lens crystallin aggregation and insolubilization. Exp Eye Res. 132:151-60. doi: 10.1016/j.exer.2015.01.024. [PubMed]