title: Amino acid residue Val362 plays a critical role in maintaining the structure of C terminus of connexin 50 and in lens epithelial-fiber differentiation.
abstract: We have previously shown that connexin (Cx) 50, unlike the other two lens connexins, Cx43 and Cx46, promotes chicken lens epithelial-fiber differentiation in a channel-independent manner. Here, we show that deletion of the PEST motif at the C terminus (CT) domain of Cx50 attenuates the stimulatory effect of Cx50 on lens fiber differentiation. Valine 362, a residue located within the PEST domain, is functionally involved. The structure of the Cx50 CT predicted by molecular modeling revealed four alpha-helices and Val(362) was found to be located in the middle of the 3rd helix. Replacement of Val(362) with amino acid residues that disrupt the alpha-helical structure predicted by molecular modeling, such as arginine, glutamate, or phenylalanine, attenuated the stimulatory effects of Cx50 on lens differentiation, whereas replacement with threonine, isoleucine, leucine, or proline, which maintain the structure preserved the function of Cx50. Circular dichroism (CD) studies supported the structural predictions and showed that the substitution with Glu, but not Thr or Pro, disrupted the alpha-helix, which appears to be the structural feature important for lens epithelial-fiber differentiation. Together, our results suggest that Val(362) is important for maintaining the helical structure and is crucial for the role of Cx50 in promoting lens epithelial-fiber differentiation.
authors.affiliation: Department of Biochemistry, University of Texas Health Science Center, San Antonio, Texas 78229-3900, USA.
authors.forename: Qian
authors.initials: Q
authors.lastname: Shi
authors.forename: Eric A
authors.initials: EA
authors.lastname: Banks
authors.forename: X Sean
authors.initials: XS
authors.lastname: Yu
authors.forename: Sumin
authors.initials: S
authors.lastname: Gu
authors.forename: Janelle
authors.initials: J
authors.lastname: Lauer
authors.forename: Gregg B
authors.initials: GB
authors.lastname: Fields
authors.forename: Jean X
authors.initials: JX
authors.lastname: Jiang
mesh_terms: D000818:Animals; D002454:Cell Differentiation; D002645:Chickens; D002942:Circular Dichroism; D017630:Connexins / Q000737:chemistry*; D004848:Epithelium / Q000473:pathology*; D005136:Eye Proteins / Q000737:chemistry*; D007908:Lens, Crystalline / Q000378:metabolism*; D018613:Microscopy, Confocal / Q000379:methods; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D014633:Valine / Q000737:chemistry*; D000099264:Connexin 50
chemical_list: D017630:Connexins; D005136:Eye Proteins; D014633:Valine; D000099264:Connexin 50
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