Significance of Interactions of Low Molecular Weight Crystallin Fragments in Lens Aging and Cataract Formation

Abstract

Analysis of aged and cataract lenses shows the presence of increased amounts of crystallin fragments in the high molecular weight aggregates of water-soluble and water-insoluble fractions. However, the significance of accumulation and interaction of low molecular weight crystallin fragments in aging and cataract development is not clearly understood. In this study, 23 low molecular mass (<3.5-kDa) peptides in the urea-soluble fractions of young, aged, and aged cataract human lenses were identified by mass spectroscopy. Two peptides, ␣B-(1-18) (MDIAIHHPWIRRPFFPFH) and ␤A3/A1-(59 -74) (SD(N)AYHIERLMSFRPIC), present in aged and cataract lens but not young lens, and a third peptide, ␥S-(167-178) (SPAVQSFRRIVE) present in all three lens groups were synthesized to study the effects of interaction of these peptides with intact ␣-, ␤-, and ␥-crystallins and alcohol dehydrogenase, a protein used in aggregation studies. Interaction of ␣B-(1-18) and ␤A3/A1-(59 -74) peptides increased the scattering of light by ␤-and ␥-crystallin and alcohol dehydrogenase. The ability of ␣-crystallin subunits to function as molecular chaperones was significantly reduced by interaction with ␣B-(1-18) and ␤A3/ A1-(59 -74) peptides, whereas ␥S peptide had no effect on chaperone-like activity of ␣-crystallin. The ␤A3/A1-(59 -74 peptide caused a 5.64-fold increase in ␣B-crystallin oligomeric mass and partial precipitation. Replacing hydrophobic residues in ␣B-(1-18) and ␤A3/A1-(59 -74) peptides abolished their ability to induce crystallin aggregation and light scattering. Our study suggests that interaction of crystallin-derived peptides with intact crystallins could be a key event in age-related protein aggregation in lens and cataractogenesis.

Spontaneous cleavage of proteins at serine and threonine is facilitated by zinc

“…Cleavage of long‐lived proteins on the N‐terminal side of Ser has been reported in proteins from a number of tissues (Table 1) (Smith et al ., 1992; Takemoto, 1995; Lampi et al ., 1998; Santhoshkumar et al ., 2008; Su et al ., 2010, 2012; Friedrich et al ., 2012) although this process has been most well documented in the lens. In the human lens, an abundant peptide is derived from the age‐related truncation of alpha B crystallin between residues 18 (His) and 19(Ser) (Santhoshkumar et al ., 2008).…”

“…In the human lens, an abundant peptide is derived from the age‐related truncation of alpha B crystallin between residues 18 (His) and 19(Ser) (Santhoshkumar et al ., 2008). To see whether this cleavage could be replicated in vitro, a peptide containing this sequence ( 16 PFHSPS 21 ) was incubated and the products characterized by mass spectrometry and NMR spectroscopy.…”

“…Its cleavage was compared with that of a Ser‐containing peptide derived from alpha A crystallin, Ac‐YEVRSDRDY. Ac‐YEVRSDRDY incorporates a sequence of alpha A crystallin (63–69) which is known to truncate at Ser 66 with age (Santhoshkumar et al ., 2008; Su et al ., 2012). As shown in Fig.…”

“…However, the mechanisms that underpin some of these processes are still poorly understood. One such modification, which is commonly observed in long‐lived proteins, is truncation at the N‐terminal side of Ser and Thr residues (Lampi et al ., 1998; Friedrich et al ., 2012, 2000; Sergeant et al ., 2003; Santhoshkumar et al ., 2008; Vlad et al ., 2011; Su et al ., 2012). In some tissues, this process could be enzymically mediated; however, proteases with specificity for cleavage next to Ser/Thr are rare.…”

“…In the adult human lens, no active proteases remain in the centre (Zhu et al ., 2010) due to the particular growth pattern of the lens, so this is an excellent tissue to examine protein cleavage processes that occur spontaneously. In this tissue, several cleavage events on the N‐terminal side of Ser and Thr have been documented (Lampi et al ., 1998; Schey et al ., 2000; Santhoshkumar et al ., 2008; Su et al ., 2012). …”