Control of amylase biosynthesis and release in the parotid gland of the rat

1. Amylase biosynthesis and release in the rat parotid were studied under various conditions. Incorporation of [(3)H]leucine into amylase, extracted from the tissue by immunoadsorbent, was measured and found to be time-dependent and totally inhibited by the protein synthesis inhibitor puromycin. 2. Adrenaline, at a concentration (10mum) that gave maximum stimulation of release, inhibited [(3)H]leucine incorporation into both total protein and amylase. This effect was reversed by phentolamine. 3. Adrenaline (1mum) and isoproterenol (10mum) stimulated biosynthesis of total protein and amylase. These effects were blocked by propranolol, as were the effects on release. Dibutyryl cyclic AMP (2mm) mimicked the effects of isoproterenol and adrenaline (1mum) on both amylase biosynthesis and release. All the above stimulatory effects on amylase biosynthesis were only observed if the tissue was pretreated with effector before pulse-labelling with [(3)H]leucine. 4. Insulin (625muunits/ml initial concentration, 150muunits/ml final concentration) stimulated incorporation of [(3)H]leucine into total protein and amylase when added to the tissue at the same time as the leucine. 5. Carbamoylcholine (10mum) decreased [(3)H]leucine incorporation into total protein and amylase when both were added to the tissue simultaneously, but this effect was prevented by removal of effector and washing the tissue before addition of [(3)H]leucine. 6. Stimulation of beta-adrenergic receptors increased both amylase release and biosynthesis, but stimulation of alpha-receptors can inhibit biosynthesis without inhibiting release. Cholinergic agents can also inhibit amylase biosynthesis, but stimulate release. Insulin at approximately physiological concentration can increase incorporation of leucine into amylase without stimulating release. The system described therefore provides an excellent model for the further investigation of the mechanisms of these diverse effects.     

Characterization of crystals of an Fab fragment of a murine monoclonal antibody.

The Fab fragment of an antibody, made against an E2-specific feline infectious peritonitis virus neutralizing antibody, has been crystallized in a form suitable for X-ray diffraction analysis from PEG 4000 using vapor diffusion methods. The Fab fragment crystals diffract to about 2.9 A resolution and are of triclinic space group P1. Unit cell dimensions, by which the reciprocal lattice can be indexed, are a = 57.16 A, b = 70.85 A, c = 75.81 A, alpha = 85.11 degrees, beta = 121.28 degrees and gamma = 116.33 degrees. There are two Fab fragments comprising the asymmetric unit of the crystals. The presence of a pseudo-mirror plane in the diffraction pattern suggests the presence of at least an approximate dyad axis relating the two Fab fragments within the asymmetric unit.     

Preliminary structure analysis of canavalin from jack bean.

The plant seed protein canavalin has been crystallized alter trypsinization by using vapor diffusion to effect isoelectric precipitation. The space groups and cell dimensions are R3 with $\text{a}\text{?}\text{= 81.3}\text{A}\text{?}\text{, γ = 111 °}$ and having extremely high R32 pseudo symmetry, P6³ with $\text{a}\text{?}\text{= 126}\text{A}\text{?}\text{and}\text{c}\text{?}\text{= 51}\text{A}\text{?}\text{, C222}{}_1\text{,}\text{with}\text{a}\text{?}\text{= 136}\text{A}\text{?}\text{,}\text{b}\text{?}\text{= 152}\text{A}\text{?}\text{, and}\text{c}\text{?}\text{= 131}\text{A}\text{?}$. X-ray data combined with electron microscopy suggests the molecule to be composed of six identical subunits. each of 18,500 daltons, arranged in a staggered hexameric ring and related by perfect or near perfect 3 2 point group symmetry. The outside diameter of the molecule is approximately 65 Å and there appears to be a large solvent channel coincident with the molecular triad.