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M Dundas

Publications and source records attributed to M Dundas.

4 recordsLinked to original sources
PubMed2001

Clinical grading of corneal staining of non-contact lens wearers.

To distinguish normal from pathological corneal fluorescein staining requires knowledge of background levels of staining among otherwise healthy individuals. Corneal staining of 102 non-contact lens wearing subjects was assessed using a photographic grading scale that uses a generic (0 to 4) scale to score corneal staining. Some degree of corneal staining was found on 79% of the corneas. Low inter-observer variability suggests that the corneal staining grading scale can be used successfully with decimal rather than integer scale increments.

Adolescent↗
PubMed1995

Heparin binding to platelet factor-4. An NMR and site-directed mutagenesis study: arginine residues are crucial for binding.

Native platelet factor-4 (PF4) is an asymmetrically associated, homo-tetrameric protein (70 residues/subunit) known for binding polysulphated glycosaminoglycans like heparin. PF4 N-terminal chimeric mutant M2 (PF4-M2), on the other hand, forms symmetric tetramers [Mayo, Roongta, Ilyina, Milius, Barker, Quinlan, La Rosa and Daly (1995) Biochemistry 34, 11399-11409] making NMR studies with this 32 kDa protein tractable. PF4-M2, moreover, binds heparin with a similar affinity to that of native PF4. NMR data presented here indicate that heparin (9000 Da cut-off) binding to PF4-M2, while not perturbing the overall structure of the protein, does perturb specific side-chain proton resonances which map to spatially related residues within a ring of positively charged side chains on the surface of tetrameric PF4-M2. Contrary to PF4-heparin binding models which centre around C-terminal alpha-helix lysines, this study indicates that a loop containing Arg-20, Arg-22, His-23 and Thr-25, as well as Lys-46 and Arg-49, are even more affected by heparin binding. Site-directed mutagenesis and heparin binding data support these NMR findings by indicating that arginines more than C-terminal lysines, are crucial to the heparin binding process.

Amino Acid Sequence↗
PubMed1993

Perturbation of the carboxy terminus of HIV-1 Rev affects multimerization on the Rev responsive element.

Perturbations within the transactivation and carboxy-terminal domains of HIV-1 Rev were examined for effects on Rev responsive element (RRE) binding activities in vitro and biological activity in vivo. Binding affinities, specificities, and multimerization of the transactivation mutants M10 and Rev/Rex M10-16 on the RRE were equivalent to wild-type Rev. Substitution of the Rex transactivation domain within Rev resulted in the incorporation of an internal methionine residue which, when cleaved with CNBr and subsequently purified, produced a protein species (CNBr-Rev) unable to fully multimerize on the RRE. Instead, two discrete protein-dependent species were generated in the gel shift assay. Furthermore, CNBr-Rev was observed to bind to the RRE with high specificity and an equilibrium binding constant of 6 x 10(-10) M. A C-terminal Rev deletion mutant (Rev M9 delta 14) lacking amino acids 68-112 displayed identical RRE binding characteristics to the CNBr-Rev protein. This protein, which lacks both the activation and the C-terminal domains, was biologically inactive but maintained the ability to discriminate the RRE from nonspecific RNA. Deletion of amino acids 92-112 resulted in a Rev mutant (Rev M11 delta 14) which bound to the RRE with wild-type affinity and high specificity. This purified mutant was observed to be aberrant in multimerization activity on the RRE with reduced multimerization apparent in the gel shift assay. However, Rev M11 delta 14 possessed biological activity equivalent to wild-type Rev in a cell-based p24 ELISA assay. These results suggest that polymerization on the RRE is dispensable for Rev activity and that two monomeric Rev proteins bound to the RRE are sufficient for biological activity. Furthermore, in vivo experiments using the Rev/Rex chimeric mutant and the M10 transdominant mutant as well as in vitro dissociation rate studies with Rev M11 delta 14 and Rev M9 delta 14 suggest that the M9 through M11 domain of the protein may be involved in RRE-dependent specific Rev dimerization.

Amino Acid Sequence↗