Biomedical subjects

A G Mackinlay

Publications and source records attributed to A G Mackinlay.

35 records · Page 2Linked to original sources

PubMed1987

Inhibition of casein and fat synthesis and alpha-lactalbumin secretion by progesterone in explants from bovine lactating mammary glands.

The effect of progesterone on casein and fat synthesis, on alpha-lactalbumin secretion and on accumulation of casein mRNA was studied in explants from bovine lactating mammary glands, cultured in the presence of insulin, cortisol or prolactin. All four biological activities were inhibited down to or below the level obtained in medium without prolactin. However, inhibition was only achieved by using concentrations of progesterone exceeding by 1000-fold those required for similar effects in tissue from non-lactating animals. A similar inhibitory effect was also found using high levels of other steroids without 11 beta-hydroxyl, such as deoxycorticosterone and dehydroepiandrosterone, and to some extent with oestradiol-17 beta, but not with cortisol or corticosterone. The inhibition by progesterone did not result from a decrease in explant viability, as determined by glucose uptake.

Animals↗

PubMed1986

Evolution of the casein multigene family: conserved sequences in the 5' flanking and exon regions.

The rat alpha- and bovine alpha s1-casein genes have been isolated and their 5' sequences determined. The rat alpha-, beta-, gamma- and bovine alpha s1-casein genes contain similar 5' exon arrangements in which the 5' noncoding, signal peptide and casein kinase phosphorylation sequences are each encoded by separate exons. These findings support the hypothesis that during evolution, the family of casein genes arose by a process involving exon recruitment followed by intragenic and intergenic duplication of a primordial gene. Several highly conserved regions in the first 200 base pairs of the 5' flanking DNA have been identified. Additional sequence homology extending up to 550 base pairs upstream of the CAP site has been found between the rat alpha- and bovine alpha s1-casein sequences. Unexpectedly, the 5' flanking promoter regions are conserved to a greater extent than both the entire mature coding and intron regions of these genes. These conserved 5' flanking sequences may contain potential cis regulatory elements which are responsible for the coordinate expression of the functionally-related casein genes during mammary gland development.

Animals↗

PubMed1984

Nucleotide sequences of bovine alpha S1- and kappa-casein cDNAs.

The nucleotide sequences corresponding to bovine alpha S1- and kappa-casein mRNAs are presented. An unusual alpha S1-casein cDNA has been characterised whose 5' end commences upstream from its putative TATA box. The alpha S1-casein mRNA is compared to rat alpha-casein mRNA and two components of divergence are identified. Firstly, the two sequences have diverged at a high point mutation rate and the rate of amino acid replacement by this mechanism is at least as great as the rate of divergence of any other part of the mRNAs. Secondly, the protein coding sequence has been subjected to several insertion/deletion events, one of which may be an example of exon shuffling . The kappa-casein mRNA sequence verifies the proposition that it has arisen from a different ancestral gene to the other caseins.

Animals↗

PubMed1983

NMR studies of the phosphoserine regions of bovine alpha s1- and beta-casein. Assignment of 31P resonances to specific phosphoserines and cation binding studied by measurement of enhancement of 1H relaxation rate.

(1) High-resolution 31P-NMR was used to study the environment of the phosphoserine residues of the phosphoproteins, alpha s1-casein B, beta-casein A2 and beta-casein C. For reference purposes 31P-NMR spectra of phosvitin and ovalbumin were also collected. (2) 31P resonances were assigned to specific phosphoserine residues as a result of comparisons of the high-resolution 31P-NMR spectra for alpha s1- and beta-caseins and for peptide fragments of these proteins obtained by cyanogen bromide and trypsin cleavage. (3) Measurements of the enhancement of the relaxation rate for water protons (1H) on addition of Mn2+ to alpha s1-casein B and to a fragment alpha s1-CN3, obtained by cyanogen bromide cleavage, gave approximate pK values for the binding groups and suggest the possibility of a conformational change induced by varying the concentration of divalent cation.

Animals↗

PubMed1982

Polynucleotide replication coupled to protein synthesis: a possible mechanism for the origin of life.

A mechanism is suggested for the replication under primitive conditions of long polynucleotides by the sequential incorporation of sequences related to those of modern transfer RNAs. It is proposed that replication of such molecules became established as the result of a replicative advantage arising from the concomitant linkage together of amino acids to form polypeptides. Initially these polypeptides may have been of random sequence. Selection of primitive tRNAs in which the amino acid and anticodon stem sequences were rotationally symmetrical could have led to specific, anticodon-directed aminoacylation and fixation of the genetic code along the lines suggested by Hopfield. (Hopfield, 1978). The primitive replication-coupled system would then have been able to synthesize specific proteins containing one amino acid residue for each primitive tRNA incorporated during replication. The end result of this line of evolution is postulated to have been a nucleoprotein structure resembling the ribosome. The primitive system would then have been able to give rise directly to triplet-coded protein synthesis. Some recent RNA sequence data are discussed which are consistent with derivation of modern protein synthesis from the primitive replication-coupled mechanism.

Amino Acid Sequence↗

PubMed1982

Construction and identification by partial nucleotide sequence analysis of bovine casein and beta-lactoglobulin cDNA clones.

Double stranded (DS) DNA molecules obtained by reverse transcription of a partially purified lactating bovine mammary gland mRNA fraction were cloned into pBR322. Restriction maps for four recombinants were constructed and partial nucleotide sequence analysis of these revealed coding sequences corresponding to alpha s1-, beta-, and kappa-casein and beta-lactoglobulin. The specific single-stranded (SS) cDNAs representing each of these species were identified and their nucleotide lengths estimated. Evidence is presented that these are essentially full-length transcripts of the major mRNA species. On this basis, the cDNA clones range in size from 50% for beta-casein to about 95% for alpha s1-casein in comparison with their respective mRNAs. The DNA sequence spanning all eight phosphoserine residues in alpha s1-casein is presented. These data, together with other serine codon usage data, indicate that the mammary gland phosphoseryl tRNA does not play a role in the incorporation of serine phosphate residues during casein synthesis. The observation that the nucleotide sequences for the serine phosphate cluster in bovine alpha s1-and rat beta-casein exhibit close homology supports the suggestion that these regions have evolved from a common primordial sequence.

Animals↗

PubMed1982

Substrate specificity studies on a calf thymus nuclear phosphoprotein kinase.

1. The protein kinase activity associated with the phosphoprotein fraction of calf thymus nuclei has been examined for its ability to phosphorylate exogenous phosphoprotein substrates. beta-Casein and phosvitin are both efficient phosphate acceptors. Phosphorylation of alpha s1 and beta-caseins was shown, directly, to occur at threonyl-49 and threonyl-41 respectively. Both threonyl residues occur within the sequence Thr-Glu-Asp. 2. alpha s1-Casein becomes a much better substrate for the kinase when partially dephosphorylated. A similar response is shown by a phosphopeptide containing the alpha s1-casein phosphate cluster and is due to a new phosphorylation site becoming available. Efficient phosphorylation of beta-casein requires that the phosphate cluster (residues 15-19) be intact and results are presented which are consistent with there being a similar requirement for phosphorylation of the site created in alpha s1-casein by partial dephosphorylation. 3. Comparison of genetic variants of beta-casein as phosphate acceptors for the kinase shows that the presence of lysyl residues close to the phosphorylation site markedly depresses the rate of phosphorylation. Maleylation of beta-casein increases the rate of phosphorylation for all of the variants tested, although to varying extents. Treatment of maleylated beta-casein with trypsin to remove the N-terminal phosphopeptide inhibits phosphorylation by the kinase. 4. The structural determinants of beta-casein allowing it to be efficiently phosphorylated by the kinase are concluded to be the presence of a sequence surrounding the phosphorylation site, which is rich in acidic amino acid residues and from which basic residues are absent. The acidic phosphate cluster of beta-casein also promotes phosphorylation either by interacting directly with the enzyme or through its influence on the conformation of beta-casein.

Animals↗

PubMed1981

Human kappa-casein as a tumor marker: purification and properties.

To test the efficacy of kappa-casein as a marker of human malignancy, a protein human milk, ostensibly kappa-casein, has been purified and serum levels determined, by RIA, in normal subjects, breast cancer patients, and lactating women. The results do not support claims by other workers for this protein. Concurrent physicochemical characterization of the protein has shown that it is probably lactoferrin and this result casts some doubt on the earlier studies as it is a major, non-beta component of the casein fraction. It also indicates that caution should be exercised when homology between proteins is used as a guide to identity.

Breast Neoplasms↗

PubMed1979

The binding of beta-casein to hydroxyapatite: the effect of phosphate content and location.

The elution behaviour of beta-casein from columns of hydroxyapatite has been studied and the effect examined of the enzymic addition of an extra phosphate residue. When the additional phosphate is located near pre-existing phosphate residues stronger binding to hydroxyapatite is observed. When the additional phosphate is remote from the pre-existing phosphates no increase in strength of binding is observed. It is suggested that the clustering of phosphate residues which characterizes alphas- and beta-caseins can be rationalized on the basis that it facilitates co-operative interactions between these phosphates and the Ca phosphate of the casein micelle and/or basic residues within casein polypeptide chains.

Amino Acid Sequence↗

PubMed1978