Expression, localization, and correlation of N-myristoyltransferase and its inhibitor in bovine eye.

Abstract

N-myristoyltransferase (NMT) is a ubiquitously distributed eukaryotic enzyme that catalyzes myristoylation of proteins. Very little is known about the process of myristoylation, particularly in the eye. In the present study, the distribution, expression, and correlation of NMT and its inhibitor (NMT inhibitor protein, NIP) in the bovine eye were investigated. Whole bovine eyes were either homogenized or regionally dissected to determine the activity and localization of NMT and NIP. Dissected tissues were homogenized, and Western blot analysis was performed using polyclonal anti-NMT and anti-NIP antibodies. The NMT activity was assayed using cAMP-dependent protein kinase or pp60(src) derived peptide as a substrate. Fresh samples were then prepared for immunohistochemical analysis and probed with polyclonal anti-NMT and anti-NIP antibodies. The total bovine eye cytosolic fraction displayed both NMT and NIP expression. NMT was present in all the regions of the eye at various levels of expression. The highest expression of NMT was in the cornea, whereas NIP was present in the retina, optic nerve, sclera, and choroid only. NIP expression was the highest in the optic nerve, sclera, and retina. NMT activity was observed in the cornea, iris, and retina after DEAE-Sepharose CL-6B column chromatography. The inhibitory activity of crude homogenate on recombinant human NMT activity was found to be greater for optic nerve and choroid. Immunohistochemistry results displayed similar findings. The varied expression of NMT in different regions of the eye reveals a regulatory relationship of NMT with NIP. These findings indicate that NMT and NIP are present in various regions of the eye and will lead to further understanding of visual signaling in ocular cells.