Faculty Bibliography

Titrations of angiotensin-converting enzyme (ACE; E.C. 3.4.15.1) present in human serum, as well as in homogenates prepared from post-mortem human caudate or mouse (C57BL1/6J) whole brain tissue, were performed with the selective ACE inhibitors, captopril (SQ 14225) and teprotide (SQ 20881). ACE activity present in human serum was more sensitive to inhibition by either inhibitor than the activity present in the brain homogenates. The inhibition curves for the titration of the human serum activity by both inhibitors were sigmoidal while the inhibition curves for the ACE activity present in the brain homogenates were more complex. These results suggest that the brain homogenates contained: at least two species of enzyme activity with properties similar to ACE but with differing affinities for the inhibitors, or substances without ACE activity that are capable of competing with ACE for the binding of the inhibitors. Therefore, measurements of captopril or teprotide-sensitive peptidase activity as well as inhibitor-binding activity may not always reflect ACE concentrations in brain tissue

Angiotensin-converting enzyme (ACE, E.C. 3.4.15.1) has been identified as a normal constituent of human cerebrospinal fluid (CSF). ACE activity in CSF from adult subjects without known neurologic disorder correlated positively (P = 0.002) with age between 50 and 90 years. Patients with moderate degrees of senile dementia of the Alzheimer's type and comparably demented patients with Parkinson's disease or progressive supranuclear palsy exhibited mean levels of ACE activity that were decreased 41, 27 and 53% respectively, compared to the mean level in an age and sex-matched group of neurologically intact individuals. These results raise the possibility that ACE activity in CSF may be an index of neuronal dysfunction in certain central neurodegenerative disorders

Cathepsin D (CD) was purified to homogeneity from postmortem human cerebral cortex. Incubation of CD with human neurofilament proteins (NFPs) prepared by axonal flotation led to the rapid degradation of the 200,000, 160,000, and 70,000 NFP subunits (200K, 160K, and 70K) which had been separated by one- or two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Degradation was appreciable at enzyme activity-to-substrate protein ratios that were two- to threefold lower than those in unfractionated homogenates from cerebral cortex. Quantitative measurements of NFPs separated by PAGE revealed that, at early stages of digestion, the 160K NFP was somewhat more rapidly degraded than the 70K subunit while the 200K NFP had an intermediate rate of degradation. At sufficiently high enzyme concentrations, all endogenous proteins in human NF preparations were susceptible to the action of CD. Human brain CD also degraded cytoskeletal proteins in NF preparations from mouse brain with a similar specificity. To identify specific NFP break-down products, antisera against each of the major NFPs were applied to nitrocellulose electroblots of NFPs separated by two-dimensional SDS-PAGE. In addition to detecting the 200K, 160K, and 70K NFP in human NF preparations, the antisera also detected nonoverlapping groups of polypeptides resembling those in NF preparations from fresh rat brain. When human NF preparations were incubated with CD, additional polypeptides were released in specific patterns from each NFP subunit. Some of the immuno-cross-reactive fragments generated from NFPs by CD comigrated on two-dimensional gels with polypeptides present in unincubated preparations. These results demonstrate that NFPs and other cytoskeletal proteins are substrates for CD. The physiological significance of these findings and the possible usefulness of analyzing protein degradation products for establishing the action of proteinases in vivo are discussed

Growing appreciation of the multiple functions of proteolytic enzymes in intracellular protein degradation and post-translational modification, in the release of biologically active macromolecules and peptides from precursors and in cellular protein regulation and quality control has stimulated interest in proteases in neurobiology and neuropathology. In this article, the proteinases and peptidases thus far studied in the human central nervous system are reviewed with respect to their enzymology, anatomical and cytological distributions and contributions to neurological and psychiatric disease states. Though information concerning brain proteases in man is fragmentary, it suffices to establish the importance of these complex systems for advancing knowledge of human cerebral function in health and disease

The use of captopril, an inhibitor of angiotensin-converting enzyme and enkephalinase, was associated with substantial mood elevation in three depressed patients. Substances that may alter neuropeptide synthesis or degradation warrant further investigation as therapeutic agents in certain neuropsychiatric disorders

Posttranslational modification of a structural protein by limited proteolysis is demonstrated for the first time in the nervous system. The 145,000 dalton subunit of neurofilaments in mouse retinal ganglion cell (RGC) axons is selectively converted in vitro to the major 143,000 and 140,000 dalton neurofilament subunits by a neutral proteinase that is activated by endogenous levels of calcium and is distinguishable from other known brain proteinases. The close similarities between this in vitro process and the previously observed modification of the 145,000 dalton neurofilament protein during axoplasmic transport in vivo suggest that the same enzymatic mechanism is involved. These findings imply that limited proteolysis is an active process along central axons in vivo and that this enzyme may play a specific role in the function of the neuronal cytoskeleton