A computationally inspired investigation of the solid forms of (R)‐1‐phenylethylammonium‐(S)‐2‐phenylbutyrate

Following the computation of a lattice energy landscape which predicted that there should be more stable, denser forms of (R)‐1‐phenylethylammonium‐(S)‐2‐phenylbutyrate, crystallizations from a range of solvents were performed to search for other polymorphs and investigate the possibility that the known P4₁ structure could be a hydrate. Extensive crystallization experiments from a wide range of solvents gave fine needles or microcrystalline samples. A redetermination of the P4₁ structure by powder X‐ray diffraction located all protons, and in conjunction with other experimental and computational evidence showed that the structure was anhydrous. Evidence for two additional forms was found as mixtures with form I. These include an orthorhombic form, possibly a Z′ = 3 polymorph, and another as yet unidentified form obtained as a minor component from dichloromethane solution. However, both these forms appear to be metastable with respect to form I (P4₁), which is therefore probably the most thermodynamically stable form that can be crystallized from solution under ambient conditions. This determination of the solid state behavior of the less readily crystallized member of the diastereomeric salt system (R)‐1‐phenylethylammonium‐(R/S)‐2‐phenylbutyrate provides a challenge to the theoretical modeling to explain its ideal resolution behavior. Chirality 2010. © 2009 Wiley‐Liss, Inc.     

High-throughput screening based identification of small molecule antagonists of integrin CD11b/CD18 ligand binding

Binding of leukocyte specific integrin CD11b/CD18 to its physiologic ligands is important for the development of normal immune response in vivo. Integrin CD11b/CD18 is also a key cellular effector of various inflammatory and autoimmune diseases. However, small molecules selectively inhibiting the function of integrin CD11b/CD18 are currently lacking. We used a newly described cell-based high-throughput screening assay to identify a number of highly potent antagonists of integrin CD11b/CD18 from chemical libraries containing >100,000 unique compounds. Computational analyses suggest that the identified compounds cluster into several different chemical classes. A number of the newly identified compounds blocked adhesion of wild-type mouse neutrophils to CD11b/CD18 ligand fibrinogen. Mapping the most active compounds against chemical fingerprints of known antagonists of related integrin CD11a/CD18 shows little structural similarity, suggesting that the newly identified compounds are novel and unique.     

Inducible Priming Phosphorylation Promotes Ligand-independent Degradation of the IFNAR1 Chain of Type I Interferon Receptor

Phosphorylation-dependent ubiquitination and ensuing down-regulation and lysosomal degradation of the interferon α/β receptor chain 1 (IFNAR1) of the receptor for Type I interferons play important roles in limiting the cellular responses to these cytokines. These events could be stimulated either by the ligands (in a Janus kinase-dependent manner) or by unfolded protein response (UPR) inducers including viral infection (in a manner dependent on the activity of pancreatic endoplasmic reticulum kinase). Both ligand-dependent and -independent pathways converge on phosphorylation of Ser⁵³⁵ within the IFNAR1 degron leading to recruitment of β-Trcp E3 ubiquitin ligase and concomitant ubiquitination and degradation. Casein kinase 1α (CK1α) was shown to directly phosphorylate Ser⁵³⁵ within the ligand-independent pathway. Yet given the constitutive activity of CK1α, it remained unclear how this pathway is stimulated by UPR. Here we report that induction of UPR promotes the phosphorylation of a proximal residue, Ser⁵³², in a pancreatic endoplasmic reticulum kinase-dependent manner. This serine serves as a priming site that promotes subsequent phosphorylation of IFNAR1 within its degron by CK1α. These events play an important role in regulating ubiquitination and degradation of IFNAR1 as well as the extent of Type I interferon signaling.     

IRAK1 and IRAK4 Promote Phosphorylation,Ubiquitination, and Degradation of MyD88 Adaptor-like (Mal)

Signal transduction by Toll-like receptor 2 (TLR2) and TLR4 requires the adaptors MyD88 and Mal (MyD88 adaptor-like) and serine/threonine kinases, interleukin-1 receptor-associated kinases IRAK1 and IRAK4. We have found that both IRAK1 and IRAK4 can directly phosphorylate Mal. In addition, co-expression of Mal with either IRAK resulted in depletion of Mal from cell lysates. This is likely to be due to Mal phosphorylation by the IRAKs because kinase-inactive forms of either IRAK had no effect. Furthermore, lipopolysaccharide stimulation resulted in ubiquitination and degradation of Mal, which was inhibited using an IRAK1/4 inhibitor or by knocking down expression of IRAK1 and IRAK4. MyD88 is not a substrate for either IRAK and did not undergo degradation. We therefore conclude that Mal is a substrate for IRAK1 and IRAK4 with phosphorylation promoting ubiquitination and degradation of Mal. This process may serve to negatively regulate signaling by TLR2 and TLR4.     

Association between adolescent idiopathic scoliosis prevalence and age at menarche in different geographic latitudes

Background

Age at menarche is considered a reliable prognostic factor for idiopathic scoliosis and varies in different geographic latitudes. Adolescent idiopathic scoliosis prevalence has also been reported to be different in various latitudes and demonstrates higher values in northern countries. A study on epidemiological reports from the literature was conducted to investigate a possible association between prevalence of adolescent idiopathic scoliosis and age at menarche among normal girls in various geographic latitudes. An attempt is also made to implicate a possible role of melatonin in the above association.

Material-methods

20 peer-reviewed published papers reporting adolescent idiopathic scoliosis prevalence and 33 peer-reviewed papers reporting age at menarche in normal girls from most geographic areas of the northern hemisphere were retrieved from the literature. The geographic latitude of each centre where a particular study was originated was documented. The statistical analysis included regression of the adolescent idiopathic scoliosis prevalence and age at menarche by latitude.

Results

The regression of prevalence of adolescent idiopathic scoliosis and age at menarche by latitude is statistically significant (p < 0.001) and are following a parallel declining course of their regression curves, especially in latitudes northern than 25 degrees.

Conclusion

Late age at menarche is parallel with higher prevalence of adolescent idiopathic scoliosis. Pubarche appears later in girls that live in northern latitudes and thus prolongs the period of spine vulnerability while other pre-existing or aetiological factors are contributing to the development of adolescent idiopathic scoliosis. A possible role of geography in the pathogenesis of idiopathic scoliosis is discussed, as it appears that latitude which differentiates the sunlight influences melatonin secretion and modifies age at menarche, which is associated to the prevalence of idiopathic scoliosis.     

"Translating" tumor hypoxia: unfolded protein response (UPR)-dependent and UPR-independent pathways

Poor oxygenation (hypoxia) is present in the majority of human tumors and is associated with poor prognosis due to the protection it affords to radiotherapy and chemotherapy. Hypoxia also elicits multiple cellular response pathways that alter gene expression and affect tumor progression, including two recently identified separate pathways that strongly suppress the rates of mRNA translation during hypoxia. The first pathway is activated extremely rapidly and is mediated by phosphorylation and inhibition of the eukaryotic initiation factor 2alpha. Phosphorylation of this factor occurs as part of a coordinated endoplasmic reticulum stress response program known as the unfolded protein response and activation of this program is required for hypoxic cell survival and tumor growth. Translation during hypoxia is also inhibited through the inactivation of a second eukaryotic initiation complex, eukaryotic initiation factor 4F. At least part of this inhibition is mediated through a Redd1 and tuberous sclerosis complex 1/2-dependent inhibition of the mammalian target of rapamycin kinase. Inhibition of mRNA translation is hypothesized to affect the cellular tolerance to hypoxia in part by promoting energy homeostasis. However, regulation of translation also results in a specific increase in the synthesis of a subset of hypoxia-induced proteins. Consequently, both arms of translational control during hypoxia influence gene expression and phenotype. These hypoxic response pathways show differential activation requirements that are dependent on the level of oxygenation and duration of hypoxia and are themselves highly dynamic. Thus, the severity and duration of hypoxia can lead to different biological and therapeutic consequences.     

Male courtship signals and female signal assessment in Photinus greeni fireflies

The evolutionary dynamic of courtship signaling systems is drivenby the interaction between male trait distributions and femalepreferences. This interaction is complex because females maychoose mates based on multiple components of male signals, andfemale preference functions may vary depending on mate availability,female reproductive state, and environmental conditions. InPhotinus fireflies (Coleoptera: Lampyridae), flying males emitbioluminescent flash signals to locate sedentary females, whichreply selectively to attractive male flash signals with theirown response flash. In this study, we first examined temporalvariation in the paired-pulse flash patterns produced by Photinusgreeni males in the field and found significant among-male variation(70% of total variation) in interpulse intervals (IPIs). Therewas no significant relationship between male IPI and spermatophoresize, suggesting that P. greeni male courtship signals do notprovide females with reliable indicators of male material resources.In laboratory playback experiments, we presented P. greeni femaleswith simulated flash signals to assess how IPI and pulse durationindependently affected the likelihood of female flash response.We also examined the effects of female body mass and time duringthe mating season on female preference functions, hypothesizingthat females would be less discriminating when they were heavier(more fecund) and when mate availability declined. We foundthat P. greeni females discriminated among signals within theirspecies' range based primarily on flash pattern IPI. Neitherthe time during the mating season nor female weight alteredfemale preference functions for IPI, although season did influencefemale response to pulse duration. These results reveal thatP. greeni females discriminate among conspecific males basedprimarily on male IPIs, the same signal character previouslyshown to be important for firefly species recognition. Fieldplayback experiments indicated that female responsiveness peakednear the average IPI given by males at different ambient temperatures,suggesting that fireflies exhibit temperature coupling similarto that seen in many acoustically signaling animals.     

An alternate conformation of the hyperthermostable HU protein from Thermotoga maritima has unexpectedly high flexibility

The homodimeric HU protein from the hyperthermophile Thermotoga maritima (HUTmar) is a model system which can yield insights into the molecular determinants of thermostability in proteins. Unusually for a thermostable protein, HUTmar exists in a structurally heterogeneous state as evidenced by the assignment of two distinct and approximately equally populated forms in solution. Relaxation measurements combined with chemical shift, hydrogen exchange, and nuclear Overhauser enhancement data confirm the main structural features of both forms. In addition, these data support a two-state model for HUTmar in which the major form closely resembles the X-ray structure while the very flexible minor form is less structured. HUTmar may therefore be a new example of the small class of hyperthermostable proteins with unexpected flexibility.