Biomedical subjects

Carolyn McKinlay

Publications and source records attributed to Carolyn McKinlay.

3 recordsLinked to original sources

PubMed2006

The Medicago truncatula small protein proteome and peptidome.

The small protein and native peptide component of plant tissues is a neglected area of proteomic studies. We have used fractionation techniques for denatured and nondenatured protein preparations combined with 2-D LC tandem mass spectrometry to examine the sequences of small proteins and peptides in four tissues of the model legume, Medicago truncatula: the root tip and root of germinating seedlings, nitrogen fixing nodules, and young leaves. The isolation and fractionation strategies successfully enriched the small protein and native peptide content of the samples. Eighty-one small M. truncatula proteins and native peptides were identified. Most samples were dominated by ribosomal and histone proteins, and leaf samples possessed photosynthesis-related proteins. Secreted proteins such as lipid transfer proteins were common to several tissues. Twenty-four hours after germination, the roots and root tip tissues possessed several "seed-specific" and late-embryogenesis proteins. We conclude that these proteins are present in cells prior to germination and that they are subsequently used as a nutritional source for the young tissues. Native UV absorbing peptides were detected in very low molecular weight fractions and sequenced. Each peptide shared C-terminal residues and showed homology to the seed storage protein legumin. The strategies used here would be suitable for combining bioassays and mass spectrometry to identify bioactive peptides in the M. truncatula peptidome.

Amino Acid Sequence↗

PubMed2004

SARS coronavirus E protein forms cation-selective ion channels.

Severe Acute Respiratory Syndrome (SARS) is caused by a novel coronavirus (SARS-CoV). Coronaviruses including SARS-CoV encode an envelope (E) protein, a small, hydrophobic membrane protein. We report that, in planar lipid bilayers, synthetic peptides corresponding to the SARS-CoV E protein forms ion channels that are more permeable to monovalent cations than to monovalent anions. Affinity-purified polyclonal antibodies recognizing the N-terminal 19 residues of SARS-CoV E protein were used to establish the specificity of channel formation by inhibiting the ion currents generated in the presence of the E protein peptides.

Amino Acid Sequence↗

PubMedDate not supplied

Synthesis of quaternised 2-aminopyrimido[4,5-d]pyrimidin-4(3H)-ones and their biological activity with dihydrofolate reductase.

In a program to design and develop mechanism-based compounds active as substrates and inhibitors of dihydrofolate reductase (DHFR), we report the synthesis and physical properties of the 6-methyl- (7), 8-methyl- (8a), and 8-ethyl- (8b) derivatives of the parent 2-aminopyrimido[4,5-d]pyrimidin-4-(3H)-one (6). These compounds are the first members of a class of heterocycles related to 8-alkylpterins (N8-alkyl-2-aminopteridin-4(8H)-ones) (2a-2c), which have been shown to be novel substrates for DHFR. Three methods were developed for the synthesis of target compounds 7, 8a and 8b; however, the optimum yields (1-8%) could not be improved because the products decomposed by ring opening (e.g. to 2,4-diamino-5-methyliminomethylpyrimidin-6(1H)-one (9)) under the reaction conditions. The marked pi-electron deficiency of compounds 7, 8a and 8b is the likely cause for the susceptibility of the quaternised pyrimidine ring in the related cations 10, 15a and 15b, respectively, to add nucleophiles, thus promoting the opening of the pyrimidopyrimidine ring system. 1H-NMR spectroscopic studies of compounds 7, 8a and 8b revealed a fast and reversible covalent hydration of the associated cations across the C7z.sbnd;N8 bond for the N6-methyl derivative 7 and across the N6z.sbnd;C7 bond for the N8-methyl derivative 8a. UV spectroscopic studies of methyl derivatives 7 and 8a as well as the parent heterocycle 6 showed that protonation of the latter occurred at N1, while methylation with iodomethane proceeded at N6 and N8. The basicities of the N-methyl derivatives 7 and 8a (pK(a) ca. 5.5) are similar to those of 8-alkylpterins 2; this is an essential element of the design to promote binding to DHFR in their protonated form. Enzyme kinetics of 7, 8a and 8b with chicken DHFR confirmed our predictions that they are substrates, with apparent K(m) values of 3.8, 0.08, and 0.65 mM, and apparent V(max) values of 0.47, 2.27, and 0.30 nmol L(-1) min(-1) (for enzyme concentration 0.122 micro M), respectively. The parent compound 6 was not a substrate.

Animals↗