No abstract

The x-ray crystal structure of recombinant human interferon-gamma has been determined with the use of multiple-isomorphous-replacement techniques. Interferon-gamma, which is dimeric in solution, crystallizes with two dimers related by a noncrystallographic twofold axis in the asymmetric unit. The protein is primarily alpha helical, with six helices in each subunit that comprise approximately 62 percent of the structure; there is no beta sheet. The dimeric structure of human interferon-gamma is stabilized by the intertwining of helices across the subunit interface with multiple intersubunit interactions.

The term "disintegrin" was first used in 1990 to describe a group of viper venom-derived, nonenzymatic small proteins that shared numerous structural and functional properties. These proteins, which have been found in a great number of viper species studied since that time possess both a remarkable sequence homology and an equally notable variability in potency and selectivity in their interactions with integrin receptors. The discovery that small disintegrins may actually have been derived from much larger mosaic proteins possessing catalytic activity, and that species other than snakes (both plant and animal) produce proteins containing disintegrin-like domains, has led to much research related to both the proteins themselves and the receptors to which they bind. The purpose of this review is to discuss the literature and the authors' own data on the structure and function of disintegrins and their relevance to the studies on proteins containing disintegrin-like domains, such as hemorrhagins and ADAMs.

The three-dimensional structure of ubiquitin has been determined at 2.8 A resolution. X-ray diffraction data for the native protein and derivatives were collected with an automated diffractometer. Phases were obtained by use of a single isomorphous mercuric acetate derivative. The molecule contains a pronounced hydrophobic core. Prominent secondary structural features include three and one-half turns of a-helix, a mixed fl-sheet that contains four strands, and seven reverse turns. The histidine, tyrosine, and two phenylalanine residues are located on the surface of the molecule. 7). It now appears that the 76-amino acid polypeptide is the physiologically active form (7,8). Ubiquitin that lacks the COOH-terminal Gly-Gly is most likely an in vitro proteolytic artifact.Ubiquitin was initially thought to be a hormone that induced lymphocyte differentiation and activated adenylate cyclase (1,9,10). However, these activities were not confirmed by later workers (11,12). More recently, interest has focused on ubiquitin-protein conjugates, in which ubiquitin is covalently linked via its carboxyl terminus to E-amino groups of lysine residues in a wide variety of intracellular proteins. The first such conjugate described was the chromosomal protein A24 (currently designated uH2A), which consists of ubiquitin conjugated to histone H2A via an isopeptide bond involving the COOH-terminal glycine of ubiquitin and the e-NH2 group of lysine-119 of histone H2A (8, 13). It was subsequently shown that ubiquitin is conjugated in vivo to all subtypes of histone H2A and also to H2B, although at much lower levels (14, 15). The function of this conjugate is not known, but it may be involved in the transcription of active genes (16).Other studies have shown that conjugation of ubiquitin with proteins is required for intracellular ATP-dependent protein degradation (17,18). The proposed mechanism of ubiquitin action in this system is through isopeptide bond formation to substrates for proteolysis; one possible role of ubiquitin is to serve as a signal for attack by proteinases specific for ubiquitin-protein conjugates. Ubiquitin-mediated, nonlysosomal proteolysis has been observed in a variety of mammalian cell types (18). Most recently, ubiquitin-protein conjugation has been studied in the mouse cell line ts85, in which the conjugation is temperature sensitive (19,20). It has been shown that this effect is due to the specific thermolability of the ts85 ubiquitin-activating enzyme. By using this cell line as a probe, it was demonstrated that the majority of short-lived proteins in this cell undergo degradation through a ubiquitin-dependent pathway. Other workers have shown that the degradation of abnormal proteins is also carried out by the ubiquitin pathway (21, 22). Thus, ubiquitin is a small protein with a large number and diversity of protein-protein interactions.Apart from its physiological roles, ubiquitin is of interest because of its stability and unparalleled sequence conservation (2-5). It is noteworthy that the molecule ...

EC3, a heterodimeric disintegrin (M r ‫؍‬ 14,762) isolated from Echis carinatus venom is a potent antagonist of ␣4 integrins. Two subunits called EC3A and EC3B were isolated from reduced and alkylated EC3 by reverse-phase high performance liquid chromatography. Each subunit contained 67 residues, including 10 cysteines, and displayed a high degree of homology to each other and to other disintegrins. EC3 inhibited adhesion of cells expressing ␣4␤1 and ␣4␤7 integrins to natural ligands vascular cell adhesion molecule 1 (VCAM-1) and mucosal addressin cell adhesion molecule 1 (MadCAM-1) with IC 50 ‫؍‬ 6 -30 nM, adhesion of K562 cells (␣5␤1) to fibronectin with IC 50 ‫؍‬ 150 nM, and adhesion of ␣IIb␤3 Chinese hamster ovary cells to fibrinogen with IC 50 ‫؍‬ 500 nM; it did not inhibit adhesion of ␣v␤3 Chinese hamster ovary cells to vitronectin. Ethylpyridylethylated EC3B inhibited adhesion of Jurkat cells to immobilized VCAM-1 (IC 50 ‫؍‬ 6 M), whereas EC3A was inactive in this system. The MLDG motif appeared to be essential for activity of EC3B. Linear MLDG peptide inhibited the adhesion of Jurkat to VCAM-1 in a dose-dependent manner (IC 50 ‫؍‬ 4 mM), whereas RGDS peptide was not active at the same concentration. MLDG partially inhibited adhesion of K562 cells to fibronectin (5-10 mM) in contrast to RGDS peptide (IC 50 ‫؍‬ 3 mM), inhibiting completely at 10 mM.