The x-ray crystal structure of recombinant human renin has been determined. Molecular dynamics techniques that included crystallographic data as a restraint were used to improve an initial model based on porcine pepsinogen. The present agreement factor for data from 8.0 to 2.5 angstroms (A) is 0.236. Some of the surface loops are poorly determined, and these disordered regions border a 30 A wide solvent channel. Comparison of renin with other aspartyl proteinases shows that, although the structural cores and active sites are highly conserved, surface residues, some of which are critical for specificity, vary greatly (up to 10A). Knowledge of the actual structure, as opposed to the use of models based on related enzymes, should facilitate the design of renin inhibitors.

No abstract

The isolation of apolipoprotein A-I-containing lipoproteins [Lp(A-I)] by selected-affinity immunosorption minimizes the loss of associated proteins that occurs during the isolation of high-density lipoproteins (HDL) by sequential ultracentrifugation. We have used two-dimensional gel electrophoretic analysis to separate the proteins associated with Lp(A-I). Using a combination of amino acid sequencing of transblotted proteins and Western blotting with specific antisera, we have identified a number of associated proteins. The positions of the apolipoproteins (apo) A-I, A-II, A-IV, C-III, D, and E were located on the gels. Lecithin-cholesterol acyltransferase and cholesteryl ester transfer protein were identified in association with Lp(A-I) to a greater extent than found associated with HDL after centrifugation. In addition to those proteins previously identified in association with HDL, we detected a number of plasma proteins associated with Lp(A-I), namely, fibrinogen, haptoglobin, proline-rich protein (C4b-binding protein), and apolipoprotein J (SP40,40 sulfated glycoprotein). The co-isolation of these proteins with Lp(A-I) does not appear to be an artifact in that they have very low affinity for a sham column containing covalently bound preimmune goat IgG in place of the anti-apoA-I IgG. These findings suggest that in addition to apolipoproteins that exist largely in association with lipoproteins there is another class of proteins which exist in lipoprotein-associated form and in the dispersed state. Detection and identification of these lipoprotein-associated proteins may aid in the mechanistic determination of a number of observed functions attributed to HDL.

No abstract

Bovine hypothalamus contains a stable, low molecular weight substance with ouabain-like properties. To further study its mechanism of action and potential physiological importance we examined its effects on purified Na+-K+-ATPase in a kinetic coupled-enzyme assay. Under optimal conditions up to 95% of Na+-K+-ATPase activity could be inhibited by the factor. Mg2+ is required for maximal inhibitory activity, but ligand requirements for optimal activity are otherwise distinct from those of both ouabain and vanadate. Inhibition is reversed by high concentrations of sodium chloride plus EDTA. Kinetic analysis yielded a Ki = 1.4 nM. The hypothalamic factor is a high-affinity reversible inhibitor of Na+-K+-ATPase, being at least as potent as the cardiac glycoside ouabain and may be a circulating inhibitor of sodium transport, which appears to be associated with experimental volume-expanded hypertension and human essential hypertension.