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Biomedical subjects

C M Henneke

Publications and source records attributed to C M Henneke.

7 recordsLinked to original sources
PubMed1995

Greek key jellyroll protein motif design: expression and characterization of a first-generation molecule.

A protein designed de novo to fold into the Greek key jellyroll structural motif has been studied. Theoretical analyses have indicated that the designed sequence should adopt the beta-strand arrangement of the Greek key jellyroll rather than any other arrangement. A synthetic gene was constructed and the protein expressed in Escherichia coli. Circular dichroism spectroscopy is consistent with the protein folding into the designed conformation and also suggests the presence of tertiary structure. Fluorescence spectroscopy showed the single tryptophan to be partially buried, while denaturation studies showed changes in fluorescence to precede alterations in secondary structure.

Amino Acid Sequence↗
PubMed1993

Protein motif by computer: the perfect Greek key jellyroll designer.

A program that generates amino acid sequences that are compatible with the Greek key protein motif is presented. Using statistical data derived from the structures of molecules from the protein databank, the novel algorithm generates amino acid sequences compatible with an 8-stranded perfect Greek key jellyroll motif. In this motif, all hydrogen bonds present in the theoretical originating beta-hairpin stay in register as the whole 8-stranded domain folds at once in an anticlockwise swirl. Eight residues are generated per strand and 32 residues per sheet making 64 residues in the antiparallel beta-barrel. The seven loops between beta-strands contain an additional 27 residues. All recognized features of beta-sheets and beta-strands, such as alternating hydrophobic, hydrophilic residues with hydrophobics on the narrow-hydrogen-bond-pair, concave side of the theoretical originating beta-ribbon; sheet twist, strand twist, side chain rotation about the strands; the theories of side chain packing between the sheets; an average 30 degrees rotation between beta-sheets; the theoretical anticomplementary patch residues of each sheet; and the anticomplementary, isotropically stressed hyperbolic parabloid shape of each sheet, are taken into account in the program. The sequences of the loops between strands are designed by turn type and strand twist is considered in the design of the motif's single beta-hairpin turn. Secondary structure parameters and between-strand amino acid pair correlations also figure importantly in the novel algorithm.

Algorithms↗
PubMed1990

Citrate synthase from the thermophilic archaebacterium Thermoplasma acidophilium. Cloning and sequencing of the gene.

The gene encoding the citric acid cycle enzyme, citrate synthase, has been cloned from the thermoacidophilic archaebacterium, Thermoplasma acidophilum. We report the sequencing of this gene and its flanking regions, and the derived amino acid sequence of the enzyme is compared by multiple-sequence alignment analysis with those of citrate synthases from eubacterial and eukaryotic organisms. The similarity is less than 30% between the archaebacterial and non-archaebacterial sequences, although the majority of residues implicated in the catalytic action of the enzyme have been conserved across all three kingdoms. The cloned archaebacterial gene has been expressed in Escherichia coli to produce catalytically active citrate synthase. This is the first reported sequence of citrate synthase from the archaebacteria.

Amino Acid Sequence↗
PubMed1989

A multiple sequence alignment algorithm for homologous proteins using secondary structure information and optionally keying alignments to functionally important sites.

The programs described herein function as part of a suite of programs designed for pairwise alignment, multiple alignment, generation of randomized sequences, production of alignment scores and a sorting routine for analysis of the alignments produced. The sequence alignment programs penalize gaps (absences of residues) within regions of protein secondary structure and have the added option of 'fingerprinting' structurally or functionally important protein-residues. The multiple alignment program is based upon the sequence alignment method of Needleman and Wunsch and the multiple alignment extension of Barton and Sternberg. Our application includes the feature of optionally weighting active site, monomer--monomer, ligand contact or other important template residues to bias the alignment toward matching these residues. A sum-score for the alignments is introduced, which is independent of gap penalties. This score more adequately reflects the character of the alignments for a given scoring matrix than the gap-penalty-dependent total score described previously in the literature. In addition, individual amino acid similarity scores at each residue position in the alignments are printed with the alignment output to enable immediate quantitative assessment of homology at key sections of the aligned chains.

Algorithms↗
PubMed1989

Sequence alignment of citrate synthase proteins using a multiple sequence alignment algorithm and multiple scoring matrices.

The alignment of Escherichia coli citrate synthase to pig heart citrate synthase and the multiple alignment of the known sequences of the citrate synthase family of enzymes have been performed using six different amino acid similarity scoring matrices and a large range of gap penalty ratios for insertions and deletions of amino acids. The alignment studies have been performed as the first step in a project aimed at homology modelling E. coli citrate synthase (a hexamer) from pig heart citrate synthase (a dimer) in a molecular modelling approach to the study of multi-subunit enzymes. The effects of several important variables in producing realistic alignments have been investigated. The difference between multiple alignment of the family of enzymes versus simple pairwise alignment of the pig heart and E. coli proteins was explored. The effects of initial separate multiple alignments of the most highly related or most homologous species of the family of enzymes upon a subsequent pairwise alignment between species was evaluated. The value of 'fingerprinting' certain residues to bias the alignment in favour of matching those residues, as well as the worth of the computerized approach compared to an intuitive alignment technique, were assessed.

Algorithms↗