A new metallothionein gene from the giant keyhole limpet Megathura crenulata

Metallothioneins (MTs) are small soluble proteins ubiquitously expressed in animals and plants. Different isoforms are present in deuterostomes and protostomes. They do not differ greatly in primary structure, but are clearly distinguishable. Here, I present the gene and the complete cDNA of a novel MT from the mollusk Megathura crenulata. This protein is closely related to the Cu-inducible MTs of the vineyard snail Helix pomatia, but has also some minor sequence features typical of Cd-inducible isoforms of H. pomatia and other molluscs. Overall, the deduced primary structure is similar to the known molluscan MTs, but in addition possesses an insertion of 5 amino acids not found in any other molluscan MTs, protostomic or deuterostomic MTs. In addition, a pentapeptide insertion, characteristic of mammalian MT-3 is present but it lacks the functional tetrapeptide CPCP within the beta-region of those MT-3 proteins that are known to suppress neuronal growth processes. The M. crenulata MT is a novel form of MT in comparison to all other known MTs. Possible functional aspects for this new MT are discussed.     

Developmental expression of two Haliotis asinina hemocyanin isoforms

Hemocyanins are large copper-containing respiratory proteins that play a role in oxygen transport in many molluscs. In some species only one hemocyanin isoform is present while in others two are expressed. The physiological relevance of these isoforms is unclear and the developmental and tissue-specific expression of hemocyanin genes is largely unknown. Here we show that two hemocyanin genes in the gastropod Haliotis asinina, which encode H. asinina hemocyanin (HaH1) and HaH2 isoforms, are developmentally expressed. These genes initially are expressed in a small number of mesenchyme cells at trochophore and pre-torsional veliger stages, with HaH1 expression slightly preceding HaH2. These cells largely are localized to the visceral mass, although a small number of cells are present in head and foot regions. Following metamorphosis the isoforms show overlapping as well as isoform-specific expression profiles, suggesting some degree of isoform-specific function.     

Human FRAG1 encodes a novel membrane-spanning protein that localizes to chromosome 11p15.5, a region of frequent loss of heterozygosity in cancer

We have previously identified a chromosomal rearrangement between fibroblast growth factor receptor 2 (FGFR2) and a novel gene, FRAG1, in a rodent model of osteosarcoma. To assess the potential role of FRAG1 in disease further, we have isolated cDNA and genomic clones of human FRAG1. Sequence analysis of the cDNA revealed the presence of an insertion not contained in the original FRAG1 sequence. This insertion in human FRAG1 encoded a region highly homologous to and immediately following the first 55 amino acids of the protein, indicating the presence of a repetitive domain within FRAG1, designated the FRAG1 homology (FH) domain. Analysis of FRAG1 gene structure revealed that the FH domains were encoded by tandem duplicated exons. Database searches identified several transmembrane proteins displaying homology to the FH domain of FRAG1. In addition, hydropathy analysis predicted FRAG1 to encode an integral membrane protein with multiple membrane-spanning segments. FRAG1 mRNA was ubiquitously expressed in human adult tissues and several tumor cell lines at varying levels of abundance. Human FRAG1 was mapped by fluorescence in situ hybridization and radiation hybrid analysis to chromosome 11 at band p15.5, a region implicated in Beckwith-Wiedemann syndrome and a region of frequent loss of heterozygosity in multiple tumor types. These results suggest that FRAG1 may be a useful candidate gene for genetic disorders associated with alterations at 11p15.5.     

Very short patch mismatch repair activity associated with gene dcm is not conferred by a plasmid coding for EcoRII methylase.

The only cytosine methylase in Escherichia coli K-12 methylates the second cytosine in the sequence CC (A/T)GG and is encoded by gene dcm. Methylation and very short patch mismatch repair activities lacking in a dcm mutant of E. coli were restored by a plasmid containing the cloned dcm gene. In contrast, plasmids with the gene for EcoRII methylase, which is a homolog of dcm, restored only cytosine methylase activity and not mismatch repair.     

Thermodynamics of anesthetic/protein interactions. Temperature studies on firefly luciferase.

Firefly luciferase is a soluble enzyme which is unusually sensitive to general anesthetics. The inhibition of the highly purified enzyme by three inhalational and three alcohol general anesthetics has been studied as a function of temperature, in the range from 5 to 20 degrees C. Inhibition constants Ki were determined at different temperatures, and van't Hoff plots of ln (Ki) versus reciprocal absolute temperature were found to be linear for all agents. Analysis of these plots gave values for the standard Gibbs free energy, enthalpy and entropy changes for transferring each anesthetic from water to the anesthetic-binding pocket on the protein. The most striking finding was that the enthalpy changes were much more negative for anesthetics binding to the protein than for binding to lipids or simple solvents. Furthermore, amongst the set of anesthetics studied, it was found that increasing potency correlated with favorable enthalpy rather than entropy changes. We discuss our results with respect to the molecular mechanisms underlying general anesthesia.     

Mapping missense and nonsense mutations in genecI of bacteriophage lambda: Marker effects

Summary Amber and missense mutations in genecI of bacteriophage lambda were mapped by reciprocal four-factor crosses, selecting recombinants between the outside markers (N amber andO amber). Distances betweencI missense mutations were additive. SeveralcIamber mutants recombined with othercI mutations with a higher-frequency than expected from the map location. Multiple exchanges in theN-O region occurred at a frequency greater than expected by chance. This high negative interference was especially marked in crosses with thecIamber mutations that were strong recombiners.A newind mutation,ind2, was found neartsU51, to the left of the previously-knownindl mutation, which is located almost in the center of genecI. The mutationc50 maps to the right oftsU50 andc71. Mutationsc60, andts71, which differ in phenotype, are apparently at the same site.