Joseph S. McGuire

Identification of the Major Postsynaptic Density Protein as Homologous with the Major Calmodulin-Binding Subunit of a Calmodulin-Dependent Protein Kinase

Abstract: The major postsynaptic density protein (mPSDp), comprising >50% of postsynaptic density (PSD) protein, is an endogenous substrate for calmodulin‐dependent phosphorylation as well as a calmodulin‐binding protein in PSD preparations. The results in this investigation indicate that mPSDp is highly homologous with the major calmodulin‐binding subunit (p) of tubulin‐associated calmodulin‐dependent kinase (TACK), and that PSD fractions also contain a protein homologous with the o‐subunit of TACK. Homologies between mPSDp and a 63,000 dalton PSD protein and the p‐ and ó‐subunits of TACK were established by the following criteria: (1) identical apparent molecular weights: (2) identical calmodulin‐binding properties; (3) manifestation of Ca2+ ‐calmodulin‐stimulated autophosphorylation; (4)identical isoelectric points; (5) identical calmodulin binding and autophosphorylation patterns on two‐dimensional gels; (6) homologous two‐dimensional tryptic peptide maps; and (7) similar phosphoamino acid‐specific phosphorylation of tubulin. The results suggest that mPSDp is a calmodulin‐binding protein involved in modulating protein kinase activity in the postsynaptic density and that a tubulin kinase system homologous with TACK exists in a membrane‐bound form in the PSD.

Activation of epidermal tyrosinase.

Abstract A soluble fraction obtained from the epidermis of Rana pipiens contains tyrosinase which either is competitively inhibited or is present as protyrosinase. Exposure of the epidermal preparation to trypsin or chymotrypsin activates the tyrosinase. There is no demonstrable activity before exposure to trypsin; after activation, dorsal (pigmented) epidermis contains roughly as much tyrosinase as ventral (white) epidermis. The activation of protyrosinase by protease offers a new site for the control of pigmentation.

Infantile acropustulosis with eosinophilic pustules

INFANTILE ACROPUSTULOSIS is a recently described clinical entit/, 2 characterized by crops of pruritic vesicopustules occurring primarily on the palms and soles. The onset is from birth to 2 months of age; the lesions may recur for up to four years. In the reported cases, all of the pustules have contained only neutrophils; this finding is said to be characteristic. On the basis of the clinical course and pathologic findings this disorder has been separated from other pustular dermatoses of infancy such as erythema toxicum neonatorum (eosinophilic infiltrate) or neonatal pustular melanosis (mixed infiltrate, predominantly neutrophilic). We are presenting a case of infantile acropustulosis with eosinophilic pustules. CASE REPORT A 5V2-month-old Caucasian boy was first seen because of recurrent crops of vcsicopustules occurring since birth. He had previously been treated with topical application of steroids and scabicides, as well as systemic administration of antibiotics and antihistamines, with no response. He had intensely pruritic pustules on the soles and sides of the feet and palms of his hands (Fig. t ). He also had lesions on his trunk and extremities. By 10 months of age the number of pustules, the frequency of appearance of new crops, and the duration of individual lesions had all decreased. At one year of age the child developed erythematous scaling patches and umbilicated firm papules diagnosed as atopic dermatitis and molluscum contagiosum, respectively. At age 17 months new crops of pruritic pustules on the hands and feet were infrequent. Smears of the pustular lesions at age 5V2 months showed many

Phosphorylation of keratin polypeptides

The phosphorylation of keratin polypeptides was examined in calf snout epidermis. When slices of epidermis were incubated in the medium containing 32Pi, the radioactivity was incorporated into several proteins. The predominant phosphorylated proteins migrated in SDS-polyacrylamide gels with apparent molecular weights between 49000 and 69000 and coincided with keratin polypeptides. The extent of keratin phosphorylation was not altered in the presence of dibutyryl cyclic AMP or reagents which elevate intracellular cyclic AMP. When homogenates of epidermis were incubated with [gamma-32P]ATP, keratin polypeptides were the predominant species phosphorylated as was also observed in epidermal slices. The presence of cyclic AMP or heat-stable inhibitor of cyclic AMP-dependent protein kinase in the reaction mixture did not affect the phosphorylation of keratin polypeptides, although the phosphorylation of exogenously-added histone was stimulated and inhibited, respectively, by these additions. Keratin polypeptides extracted from calf snout epidermis by 8 M urea were phosphorylated by incubation with [gamma-32P]ATP and cyclic AMP-dependent protein kinase from calf snout epidermis or bovine heart. No proteins were phosphorylated without the addition of the enzymes. The presence of cyclic AMP in the reaction mixture stimulated the keratin phosphorylation, and further addition of heat-stable protein kinase inhibitor reduced this stimulation.

Coupled aldehyde dehydrogenase and catalase in mitochondrial extracts

Abstract Nicotinamide adenine dinucleotide is reduced to NADH 2 in extracts of mitochondria in the presence of ethanol and cysteine. The generation of NADH 2 is dependent on two coupled reactions in mitochondrial extracts: the oxidation of ethanol to acetaldehyde by the peroxidatic action of catalase, and the further oxidation of aldehyde by aldehyde dehydrogenase. At the present, no physiological role can be assigned to this couple; however, it constitutes an avid aldehyde trap and generator of NADH 2 .

The Mechanism of Action of Melanocyte Stimulating Hormone

The mechanism of hormonal action is a central problem of biochemistry and physiology. There have been in the past few years promising attempts at its dissolution especially in the case of ACTH and the steroids, and there are hypotheses which may eventually be found to be correct. If by „the mechanism of action“ one means the initial unique reaction involving the hormone which ultimately accounts for its physiological effect, then at the present time we are unable to offer a completely satisfactory explanation for the mechanism of action of a single hormone. Before discussing current studies with MSH, I would like to discuss briefly the study of pigmentation as an area of interest to endocrinologists.

THE MECHANISM OF ACTION OF THE MELANOCYTE STIMULATING HORMONES

The melanocyte stimulating hormones, \g=a\-and \g=b\-MSH,are found in the pituitary glands of mammals. These substances are linear polypeptides composed of 13 to 22 amino acids. In very small concentration they can darken the melanocytes of frogs and some other marine animals. The metabolic role of these hormones in man and other mammals is unknown. However, there is good evidence that MSH, when released by the pituitary gland in excessive amounts as in patients with adrenal cortical insufficiency, produces hyperpigmentation of the skin. Thus far only a single \g=a\-MSHbut 3 \g=b\-MSHshave been found. \g=a\-MSHfrom hogs, cattle and sheep is made up of 13 amino acids which are the same as the first 13 amino acids of corticotrophin (ACTH). However, the N terminal serine has an N-acetyl group and the C terminal valine is in the form of an amide. Recently \g=a\-MSHwas found in human and monkey pituitary glands. The structure of \g=a\-MSHfrom human and monkey sources will be described if current investigations are completed before the meetings. Experiments on the isolation and determination of the structure of/3-MSH from human and monkey pituitary glands also will be presented. The synthesis of a-MSH by K. Hofmann and collaborators has made avail¬ able many intermediate peptides ranging from 5 to 13 amino acids as by-pro¬ ducts. Most of the small peptides can darken frog skin. Darkening activity increases as the peptide structure becomes more similar to that of natural a-MSH. The activity of synthetic a-MSH with and without various blocking groups will be discussed. Synthetic and natural a-MSH, porcine ß-MSH and porcine ACTH will be given to a human subject at different times. The skin darkening effects of these four peptides after parenteral administration to human beings will be compared. Department of Medicine, Yale University School of Medicine, New Haven, Conn.