A two-base mechanism by which the histidine and aspartic acid together catalyze dehydration and isomerization reactions is consistent with the active-site structure. The unique topology of the protein fold and the identification of the active-site components reveal features of predictive value for another enzyme, FabZ, which may be the non-specific dehydratase involved in elongation of fatty acyl chains. A positively charged area surrounding the entrance to the active site, which could interact with the negatively charged ACP, was also found.

Proteins with expanded polyglutamine domains cause eight inherited neurodegenerative diseases, including Huntington's, but the molecular mechanism(s) responsible for neuronal degeneration are not yet established. Expanded polyglutamine domain proteins possess properties that distinguish them from the same proteins with shorter glutamine repeats. Unlike proteins with short polyglutamine domains, proteins with expanded polyglutamine domains display unique protein interactions, form intracellular aggregates, and adopt a novel conformation that can be recognized by monoclonal antibodies. Any of these polyglutamine length-dependent properties could be responsible for the pathogenic effects of expanded polyglutamine proteins. To identify peptides that interfere with pathogenic polyglutamine interactions, we screened a combinatorial peptide library expressed on M13 phage pIII protein to identify peptides that preferentially bind pathologic-length polyglutamine domains. We identified six tryptophan-rich peptides that preferentially bind pathologic-length polyglutamine domain proteins. Polyglutamine-binding peptide 1 (QBP1) potently inhibits polyglutamine protein aggregation in an in vitro assay, while a scrambled sequence has no effect on aggregation. QBP1 and a tandem repeat of QBP1 also inhibit aggregation of polyglutamine-yellow fluorescent fusion protein in transfected COS-7 cells. Expression of QBP1 potently inhibits polyglutamine-induced cell death. Selective inhibition of pathologic interactions of expanded polyglutamine domains with themselves or other proteins may be a useful strategy for preventing disease onset or for slowing progression of the polyglutamine repeat diseases.Eight inherited neurodegenerative diseases, including Huntington's disease, dentatorubral pallidoluysian atrophy, spinobulbar muscular atrophy, and spinocerebellar ataxia types 1, 2, 3, 6 and 7, are caused by expanded CAG repeats in the coding region of the disease genes (1-3). The CAG codon is translated into glutamine, and the polyglutamine domain is the only region of homology among the eight disease proteins. The length of the repeat is the critical determinant of age-of-disease onset, with repeat length greater than 40 glutamines producing neurodegeneration in seven of the eight diseases (1-3).Proteins with pathologic-length polyglutamine domains display novel properties that are not present in these proteins when they contain a shorter polyglutamine domain. Length-dependent polyglutamine-protein interactions are reported for Huntington-associated protein 1, glyceraldehyde-3-phosphate dehydrogenase, leucine-rich acidic nuclear protein, vimentin, neurofilament, apopain, calmodulin, WW domain proteins, and Ras-related nuclear protein/ARA24 (4 -12). Proteins with expanded polyglutamine domains also aggregate, and aggregation is a pathologic hallmark of the polyglutamine repeat diseases (13,14). These polyglutamine length-dependent properties may arise from the ability of long polyglutamine domains to adopt unique three-dimensional confor...

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The genetic code is based on aminoacylation reactions where specific amino acids are attached to tRNAs bearing anticodon trinucleotides. However, the anticodonindependent specific aminoacylation of RNA minihelix substrates by bacterial and yeast tRNA synthetases suggested an operational RNA code for amino acids whereby specific RNA sequences/structures in tRNA acceptor stems correspond to specific amino acids. Because of the possible significance of the operational RNA code for the development of the genetic code, we investigated aminoacylation of synthetic RNA minihelices with a human enzyme to understand the sequences needed for that aminoacylation compared with those needed for a microbial system. We show here that the species-specific aminoacylation of glycine tRNAs is recapitulated by a speciesspecific aminoacylation of minihelices. Although the mammalian and Escherichia coli minihelices differ at 6 of 12 base pairs, two of the three nucleotides essential for aminoacylation by the E. coli enzyme are conserved in the mammalian minihelix. The two conserved nucleotides were shown to be also important for aminoacylation of the mammalian minihelix by the human enzyme. A simple interchange of the differing nucleotide enabled the human enzyme to now charge the bacterial substrate and not the mammalian minihelix. Conversely, this interchange made the bacterial enzyme specific for the mammalian substrate. Thus, the positional locations (if not the actual nucleotides) for the operational RNA code for glycine appear conserved from bacteria to mammals.The 20 synthetases are divided into two classes (I and II) of 10 enzymes each (1, 2). Enzymes of each class are approximately comprised of two major domains, where each of the two tRNA domains interacts with a distinct domain of the cognate tRNA synthetase (3-5). The tRNA acceptor-TTC helix ("minihelix" domain) interacts with the class-defining catalytic domain (and insertions into that domain), and segments outside of the acceptor-TTC stem-loop, such as the anticodon, interact with a second, highly variable synthetase domain that is joined to the catalytic core. While many synthetases make contact with their tRNA anticodons, specific mutations in the acceptor stems of their tRNAs severely reduce aminoacylation efficiency (6-8) and, even for these "anticodon" examples, RNA oligonucleotides with sequences based on acceptor stems alone are also aminoacylated with their cognate amino acids (9-11). The operational RNA code based on acceptor stems may have predated the genetic code and was possibly incorporated into or combined with the genetic code when the two domains of tRNAs were assembled into a single molecule (4).Escherichia coli glycyl-tRNA synthetase has an a212 quaternary structure, with a 303-amino acid a chain and 689-amino acid 13 subunit (12, 13). In contrast, the human enzyme is an a2 dimer of 739-amino acid polypeptides (14, 15), similarThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be her...

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