In this research, the necessity of understanding and using bioinformatics is demonstrated using the enzyme aspartate transcarbamoylase (ATCase) as the model enzyme. The first portion of this research focuses on the use of bioinformatics. A partial sequence of the pyrB gene found in Enterococcus faecalis was submitted to GenBank and was analyzed against the contiguous sequence from its own genome project. A BLAST (Basic Local Alignment Search Tool; Atschul, et al., 1990) was performed in order to hypothesize the remaining portion of the gene from the contiguous sequence. This allowed a global comparison to other known aspartate transcarbamoylases (ATCases) and once deduced, a translation of the sequence gave the stop codon and thus the complete sequence of the open reading frame. When this was complete, upstream and downstream primers were designed in order to amplify the gene from genomic DNA. The amplified product was then sequenced and used later in phylogenetic analyses concerning the evolution of ATCase. The second portion of this research involves taking multiple ATCase nucleotide sequences and performing phenetic and phylogenetic analyses of the archaea and eubacter families. From these analyses, ancestral relationships which dictate both structure and function were extrapolated from the data and discussed.

Signal transduction via the phosphoinositide (PI) second messenger system has key roles in the development and plasticity of the neocortex. The present study localized PI responses to individual cortical layers in slices of developing rat somatosensory cortex. The acetylcholine agonist carbachol and the glutamate agonist trans-1-amino-1,3-cyclopentanedicarboxylic acid (ACPD) were used to stimulate PI turnover. The PI responses were compared to the distribution of the corresponding PI-linked receptors in order to investigate the regional ontogeny of PI coupling to receptors in relation to neural development. The method for assessing PI turnover was modified from Hwang et al. (1990). This method images the PI response autoradiographically through the localizaton of [3H]cytidine that has been incorporated into the membrane-bound intermediate, cytidine diphosphate diacylglycerol. In each age group (postnatal days 4-30), carbachol resulted in more overall labeling than ACPD. For both agonists, the response peaked on postnatal day 10 (P10) and was lowest in the oldest age group. The laminar distribution of the carbachol PI response from P4-P16 corresponded fairly well with the laminar distribution of [3H]quinuclidinyl benzilate binding (Fuchs, 1995). However, in the subplate layer the carbachol response was strong while receptor binding was minimal. The carbachol response decreased after postnatal day 10, …