Mono-ADP-ribosylation is a major post-translational modification performed by bacterial toxins, which transfer an ADP-ribose moiety to a substrate acceptor residue. Actin- and Rho-specific ADP-ribosylating toxins (ARTs) are typical ARTs known to have very similar tertiary structures but totally different targets. Actin-specific ARTs are the A components of binary toxins, ADP-ribosylate actin at Arg177, leading to the depolymerization of the actin cytoskeleton. On the other hand, C3-like exoenzymes are Rho-specific ARTs, ADP-ribosylate Rho GTPases at Asn41, exerting an indirect effect on the actin cytoskeleton. This review focuses on the differences and similarities of actin- and Rho-specific ARTs, especially with respect to their substrate recognition and cell entry mechanisms, based on structural studies.

Unusual outbreaks of food poisoning in Japan were reported in which Clostridium perfringens was strongly suspected to be the cause based on epidemiological information and fingerprinting of isolates. The isolated strains lack the typical C. perfringens enterotoxin (CPE) but secrete a new enterotoxin consisting of two components: C. perfringens iota-like enterotoxin-a (CPILE-a), which acts as an enzymatic ADP-ribosyltransferase, and CPILE-b, a membrane binding component. Here we present the crystal structures of apo-CPILE-a, NAD+-CPILE-a and NADH-CPILE-a. Though CPILE-a structure has high similarity with known iota toxin-a (Ia) with NAD+, it possesses two extra-long protruding loops from G262-S269 and E402-K408 that are distinct from Ia. Based on the Ia–actin complex structure, we focused on actin-binding interface regions (I-V) including two protruding loops (PT) and examined how mutations in these regions affect the ADP-ribosylation activity of CPILE-a. Though some site-directed mutagenesis studies have already been conducted on the actin binding site of Ia, in the present study, mutagenesis studies were conducted against both α- and β/γ-actin in CPILE-a and Ia. Interestingly, CPILE-a ADP-ribosylates both α- and β/γ-actin, but its sensitivity towards β/γ-actin is 36% compared with α-actin. Our results contrast to that only C2-I ADP-ribosylates β/γ-actin. We also showed that PT-I and two convex-concave interactions in CPILE-a are important for actin binding. The current study is the first detailed analysis of site-directed mutagenesis in the actin binding region of Ia and CPILE-a against both α- and β/γ-actin.

Estrogen receptors (ERs) are members of 7-transmembrane receptors such as steroid hormone receptor subfamily, G protein coupled receptor family, and nuclear hormone receptor superfamily. The amino acid sequences are different in types and depend on species (Kumar and Thompson, 1999;Kumar et al., 2011). There are two subtypes of estrogen receptor; ERα and ERβ (Kuiper et al., 1997;Barkhem et al., 1998). The genes encoding ERs located on different chromosomes, which are species specific. For example, ERα locates on chromosome 6th and ERβ on chromosome 14th in humans. In mice, ERα are on the 10th and ERβ on the 12th whereas ERα locates on the 1st and ERβ on the 6th in rats. ERα are on the 1st and ERβ on the 2nd in dog comparison to ERα on B2 and ERβ on B3 in cat.ERs consist of 5 domains; 1) N-terminal domain (NTD), 2) DNA binding domain (DBD), 3) Hinge region, 4) Ligand binding domain (LBD), and 5) Agonistantagonist distinct (C-terminal domain) (Lewis et al., 2002;Kumar et al., 2011). ERs-ligands interaction are attributed to changing LBD conformation. The binding affinity is calculated by measurement of the strength of the interaction between LBD and such ligands via

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Hormone-related mammary gland tumors are among the most commonly diagnosed neoplasms in female dogs. Estrogen enacts its biological roles through specific receptors known as estrogen receptors (ER). In human medicine, anti-estrogen therapy has become the gold standard in ER-positive breast tumors' therapeutic regimen. The binding pocket of the canine estrogen receptor alpha (cERα) ligand binding domain comprises of three key amino acid residues including E354, G522 and L526, which stabilize the cERα-E2 interaction via hydrogen bonding. The side chain of E354 shares hydrogen bond interaction with the A ring of its natural ligand E2, whereas the main chain of G522 and L526 interact with the E2-D ring. The single mutation of the E354 aberrant, along with the hydrogen bond interaction between cERα and both ligands, leads to a variety of binding affinities. According to this in silico model, it may be concluded that E354 plays a role in the cERα activities. The effects of single mutants might need to be studied further in vitro and in vivo.