Studies of membrane association of CLN3 protein

The product of the CLN3 gene is a novel protein of unknown function. Simula tions using amphiphacy algorithms have shown that structurally CLN3 may be another candidate for the family of membranous proteins. Signals controllin g intracellular targeting of many membrane proteins are present as short se quences within their cytoplasmic domains. In fact, the sequence of CLN3 pro tein contains several such signaling sequences, which are conserved among m ammals. First, at the N-terminus, potential N-myristoylation motif is prese nt. Second, the C-terminal part of CLN3 protein contains both the dileucine motif, which is a potential lysosomal targeting signal, and the prenylatio n motif. There is scanty evidence of lysosomal and/or mitochondrial localiz ation of CLN3 protein. However, the question of where the functional site o f the cln3 protein exists in vivo remains unanswered. From theoretical calc ulations, we hypothesized that CLN3 should be an integral part of the membr anous micro-environment. First, to test this hypothesis, we initiated deter gent-partitioning experiments, localizing CLN3 predominantly in a pool of m embranous protein. Further studies have shown that CLN3 protein integrates spontaneously with cellular membranes. Second, based on the prenylation res ults of CLN3 protein in vitro, we discussed the possible topological conseq uences of C-terminal fragment of CLN3 protein. (C) 1999 Academic Press.