The potentiation of Ca(v)2.2 currents by microinjection of PKC beta II and PKC epsilon isozymes was not altered by the inhibition of G proteins with GDP beta S. The combination of isozyme specific inhibitors with previously generated Ser/Thr to Ala mutants of alpha(1) 2.2 subunit revealed that PKC beta II or PKC epsilon isozymes (but not PKC alpha or delta) can provide full enhancement through

the stimulatory site (Thr-422) in the I-II linker but that PKC epsilon is better at decreasing channel activity through the inhibitory site Ser-425. The enhancing effect of PKC beta II or epsilon at Thr-422 is dominant over the inhibitory effect at Ser-425. Injected PKC beta II also enhances Ca(v)2.2 current when any of the potential stimulatory sites (Ser-1757, Ser-2108

and Ser-2132) are available in the C-terminus. PKC epsilon provides lesser enhancement with C-terminal sites and only with Ser-2108 and Ser-2132. Sites GSK461364 clinical trial Ser-1757 and Ser-2132, but not Ser-2108, are dominant over the inhibitory site Ser-425. Collectively, these results reveal a hierarchy of regulatory sites in Ca(v)2.2 channels. Site-specific regulation by different PKC isozymes may allow graded levels of channel activation and susceptibility buy Cl-amidine or resistance to subsequent stimulatory events. (C) 2009 IBRO. Published by Elsevier Ltd. All rights reserved.”
“Earlier studies showed that human lens ALDH1A1 plays a critical role in protection against oxidative stress-induced cytotoxicity in human lens epithelial cells (HLEC), and opacification of rat and mouse lens. The complete coding sequence of ALDH1A1 was cloned from human lens cDNA library by using PCR methods and expressed it in Escherichia coli. The cloned human lens ALDH1A1 cDNA encodes a 501-amino-acid protein (molecular mass = 54.8 kD) that is 100% identical to human liver ALDH1A1 and shares significant identity with the same isozyme from other tissues

and species. The purified recombinant human lens ALDH1A1 exhibited optimal Exoribonuclease catalytic activity at pH 8 and preferred NAD+ as cofactor and specifically catalyzed the oxidation of toxic lipid aldehydes such as 4-hydroxynonenal (HNE; Km = 4.8 M) and malonaldehyde (Km MDA = 3.5 M). Citral, disulfiram, and cyanamide were found to inhibit human lens ALDH1A1 at IC50 values of 55, 101, and 22610 M, respectively, whereas diethylstilbestrol (DES) was found to be an activator (EC50, 1.3 M). Further, modification of recombinant human lens ALDH1A1 with nitric oxide donors such as S-nitroso-N-acetylpenicillamine (SNAP) and S-nitrosoglutathione (GSNO) significantly inhibited the enzyme activity. It therefore appears that activation of ALDH1A1, which efficiently catalyzes the detoxification of lipid-derived toxic aldehydes, and/or prevention of its oxidative modification may be novel therapeutic interventions against oxidative stress-induced lens pathologies.