Triose phosphate isomerase, a novel enzyme‐crystallin, and τ‐crystallin in crocodile cornea

Abstract

Crystallins are defined as lens structural proteins, which are classified as ubiquitous a-, b-and c-crystallins, and taxon-specific crystallins [1]. The transparency and refractive properties of lens depends on crystallins. Taxon-specific crystallins are generally enzymes recruited as structural proteins in the lens to perform specialized functions, i.e., maintaining lens transparency [2]. Some examples of the species-specific crystallins are d-and e-crystallins present in birds and reptiles, s-crystallins in turtle and reptiles, q-crystallins in frog and camel lenses [3]. Several of these crystallins are metabolic enzymes: d-crystallin is argininosuccinate lyase [4]; s-crystallin is a-enolase [5]. They are believed to be recruited by gene sharing or by gene duplication [2,6].Some enzymes such as aldehyde dehydrogenase class 3 (ALDH3), aldehyde dehydrogenase class 1 Several enzymes are known to accumulate in the cornea in unusually high concentrations. Based on the analogy with lens crystallins, these enzymes are called corneal crystallins, which are diverse and species-specific. Examining crystallins in lens and cornea in multiple species provides great insight into their evolution. We report data on major proteins present in the crocodile cornea, an evolutionarily distant taxon. We demonstrate that s-crystallin ⁄ a-enolase and triose phosphate isomerase (TIM) are among the major proteins expressed in the crocodile cornea as resolved by 2D gel electrophoresis and identified by MALDI-TOF. These proteins might be classified as putative corneal crystallins. s-Crystallin, known to be present in turtle and crocodile lens, has earlier been identified in chicken and bovine cornea, whereas TIM has not been identified in the cornea of any species. Immunostaining showed that s-crystallin and TIM are concentrated largely in the corneal epithelium. Using western blot, immunofluorescence and enzymatic activity, we demonstrate that high accumulation of s-crystallin and TIM starts in the late embryonic development (after the 24th stage of embryonic development) with maximum expression in a two-week posthatched animal. The crocodile corneal extract exhibits significant a-enolase and TIM activities, which increases in the corneal extract with development. Our results establishing the presence of s-crystallin in crocodile, in conjunction with similar reports for other species, suggest that it is a widely prevalent corneal crystallin. Identification of TIM in the crocodile cornea reported here adds to the growing list of corneal crystallins.Abbreviations ALDH, aldehyde dehydrogenase; IEF, isoelectric focusing; TIM, triose phosphate isomerase.

Corneal crystallins and the development of cellular transparency

“…Since the corneal epithelium and the lens are both derived from ectoderm, it seems reasonable to propose that the evolution of transparency in these two structures may have co-evolved as an integrated 'refracton' unit. Indeed, data showing that the corneas from a diverse range of species including squid, fish, frogs, crocodiles, birds and mammals abundantly express a few water-soluble proteins is consistent with this theory [6,7,[9][10][11]13]. However, the vast majority of studies identifying corneal crystallin expression have used total corneal extracts or isolated corneal epithelium.…”

“…Later studies have identified BCP54 as aldehyde dehydrogenase 3A1 [4,5] that comprises from 20-40% of the total water-soluble protein content of the bovine corneal epithelium. More recent studies have shown that unlike non-transparent tissues and organs other than lens, the corneas from a wide range of species abundantly express a few water-soluble enzyme/proteins (Table 1) [3,[6][7][8][9][10][11][12][13][14], many of which are identical to the abundantly expressed taxon-specific lens crystallins, including aldehyde dehydrogenase 1A1 (ALDH1A1)/η-crystallin, α-enolase/τ-crystallin, glutathione-S-transferase/Ω-crystallin, lactic dehydrogenase/ε-crystallin, glyceraldehyde-3-phosphate dehydrogenase (G3PDH)/π-crystallin, and arginino-succinate lyase/δ-crystallin [6,8]. Interestingly, several of these corneal enzyme/proteins are abundantly expressed in the lens of the same species, notably argininosuccinate lyase/δ-crystallin in the chicken and glutathione-S-transferase/Ω-crystallin in the squid.…”

Ubiquitous Lens α-, β-, and γ-Crystallins Accumulate in Anuran Cornea as Corneal Crystallins

“…Corneal epithelium of these species is known to accumulate several metabolic enzymes, such as aldehyde dehydrogenase (7-9), ␣-enolase (6,13,14), or gelsolin (12). In this study, we have selected the Indian toad as a representative of anuran amphibian and identified major soluble proteins in the cornea.…”

“…For example, aldehyde dehydrogenases 1 and 3 (7-9), transketolase (10), and isocitrate dehydrogenase (11) are expressed in human and bovine cornea, cyclophilin in the chicken cornea (6), and gelsolin in the zebrafish cornea (12), whereas ␣-enolase is expressed in the corneas of many species (6,13,14). Because their expression in corneal epithelium is significantly high, these proteins have been termed corneal crystallins, analogous to the multifunctional lens crystallins (9,(15)(16)(17).…”

“…Although not many examples of crystallins common to the lens and cornea are known, there are a few examples such as -crystallin, which is present in the lenses of several species and is also a corneal crystallin (6,14). Some invertebrates (e.g.…”