Showing papers in "Advances in Enzymology and Related Areas of Molecular Biology in 2006"

Solid-phase peptide synthesis.

Abstract: It is a privilege to be able to contribute to this volume in which Professor Zervas’ friends, students, and colleagues have joined together to honor him and his remarkable contributions to peptide chemistry. Although I did not know him until after his days in the Bergmann Laboratory at the Rockefeller Institute, I can claim the distinction of now working in those very same rooms that they occupied back in the mid-1930s. Like all peptide chemists, I am greatly indebted to Professor Zervas, having depended so heavily on the carbobenzoxy group and on the various modified urethan protecting groups which have been direct extensions of the revolutionary advance that Bergmann and Zervas made.

Cysteinyl proteinases and their selective inactivation.

Abstract: The affinity-labeling of cysteinyl proteinases may now be carried out with a number of peptide-derived reagents with selectivity, particularly for reactions carried out in vitro. These reagents have been described with emphasis on their selectivity for cysteine proteinases and lack of action on serine proteinases, the most likely source of side reactions among proteinases. Perhaps a crucial feature of this selectivity is an enzyme-promoted activation due to initial formation of a hemiketal, which may destabilize the reagent. Prominent among the reagent types that have this class selectivity are the peptidyl diazomethyl ketones, the acyloxymethyl ketones, the peptidylmethyl sulfonium salts, and peptidyl oxides analogous to E-64. The need for specific inhibitors capable of inactivating the target enzyme in intact cells and animals is inevitably pushing the biochemical application of these inhibitors into more complex molecular environments where the possibilities of competing reactions are greatly increased. In dealing with the current state and potential developments for the in vivo use of affinity-labeling reagents of cysteine proteinases, the presently known variety of cysteinyl proteinases had to be considered. Therefore this chapter has, at the same time, attempted to survey these proteinases with respect to specificity and gene family. The continual discovery of new proteinases will increase the complexity of this picture. At present the lysosomal cysteine proteinases cathepsins B and L and the cytoplasmic calcium-dependent proteinases are reasonable goals for a fairly complete metabolic clarification. The ability of investigators to inactivate individual members of this family in vivo, possibly without complications due to concurrent inactivation of serine proteinases by improvements in reagent specificity, is increasing. Among the cysteine proteinases, at least those of the papain super family, hydrophobic interactions in the S2 and S3 subsites are important and some specificity has been achieved by taking advantage of topographical differences among members of this group. Some of this has probably involved surface differences removed from the regions involved in proteolytic action. The emerging cysteine proteinases include some which, in contrast to the papain family, have a pronounced specificity in S1 for the binding of basic side chains, familiar in the trypsin family of serine proteinases. At least a potential conflict with serine proteinases can be avoided by choice of a covalent bonding mechanism. The departing group region, has not been exploited. As a sole contributor to binding, this region may be rather limited as a source of specificity.(ABSTRACT TRUNCATED AT 400 WORDS)

Ceruloplasmin: the copper transport protein with essential oxidase activity.

Abstract: Ceruloplasmin, the blue copper-protein of vertebrate plasma, has been reviewed mainly from a functional point of view. However we have surveyed the chemistry and state copper in the molecule because of the implications of the recent data of Ryden (13,28). His observations suggest that unless special precautions are taken in the isolation of ceruloplasmin degradation, probably proteolytic, produces fragments of various sizes. When isolated, these fragments appear to be held together by noncovalent interactions. Comparison of their catalytic and spectral properties reveals no significant differences from a single homogeneous species of molecular weight of 134,000 isolated by Ryden's methods. On the other hand, the homogeneous molecule may differ in properties highly sensitive to conformation and three-dimensional parameters. Three types of copper atoms have been identified in ceruloplasmin, but their amino acid environment is still unknown. Ceruloplasmin possesses significant oxidase activity towards Fe(II) and numerous aromatic amines and phenols. Its ferroxidase activity has led to the discovery that it is a molecular link between copper and iron metabolism. Ceruloplasmin mobilizes iron into the plasma from iron storage cells in the liver. An equally important duty is that ceruloplasmin, after its rapid biosynthesis in the liver, serves as a major copper transport vehicle, comparable to transferrin. Evidence is accumulating that the copper atoms of ceruloplasmin are a prerequisite for copper utilization in the biosynthesis of cytochrome oxidase and other copper proteins. The ability of ceruloplasmin to release copper at specific cellular sites may be related to its broad substrate spectrum of biological reducing agents. A possible third role of ceruloplasmin is as a contributor to the regulation of the balance of biogenic amines through its oxidase action on the epinephrine and the hydroxyindole series. Thus ceruloplasmin is a copper-protein with several important functions, all of which are directly related to its oxidase activity.