Polymorphic forms of nucleic acids provide platforms for new nanomaterials, and transition metal cations give access to alternative arrangements of nucleobases by coordinating with electron-rich functional groups. Interaction of Ag+ with 5’-guanosine monophosphate (5’-GMP) is considered in this work. Ag+ promotes nucleotide stacking and aggregation, as indicated by the increased viscosity of 5’-GMP solutions with Ag+, magnification of the circular dichroism response of guanine by Ag+, and exothermic reactions between Ag+ and guanine derivatives. Isothermal titration calorimetry studies show that the reaction is favored starting at 10 μM 5’-GMP. Utilizing the exothermic heat change associated with reaction of Ag+ with 5’-GMP, local structure within the aggregate was assessed. Based on salt dependence and comparison with the corresponding nucleoside, the dianionic phosphate of 5’-GMP is one binding site for Ag+, although this electrostatic interaction is not a dominant contribution to the overall heat change. Another binding site is the N7 on the nucleobase, as determined via studies with 7-deazaguanosine. Besides this binding site, Ag+ also associates with the O6, as earlier studies deduced from the shift in the carbonyl stretching frequency associated with adduct formation. With these two binding sites on the nucleobase, the empirical stoichiometry of ~1 Ag+:nucleobase derived from the calorimetry studies indicates that Ag+ coordinates two nucleobases. The proposed structural model is a Ag+-mediated guanine dimer within a base stacked aggregate.

The indole-diterpene paxilline is an abundant secondary metabolite synthesized by Penicillium paxilli. In total, 21 genes have been identified at the PAX locus of which six have been previously confirmed to have a functional role in paxilline biosynthesis. A combination of bioinformatics, gene expression and targeted gene replacement analyses were used to define the boundaries of the PAX gene cluster. Targeted gene replacement identified seven genes, paxG, paxA, paxM, paxB, paxC, paxP and paxQ that were all required for paxilline production, with one additional gene, paxD, required for regular prenylation of the indole ring post paxilline synthesis. The two putative transcription factors, PP104 and PP105, were not co-regulated with the pax genes and based on targeted gene replacement, including the double knockout, did not have a role in paxilline production. The relationship of indole dimethylallyl transferases involved in prenylation of indole-diterpenes such as paxilline or lolitrem B, can be found as two disparate clades, not supported by prenylation type (e.g., regular or reverse). This paper provides insight into the P. paxilli indole-diterpene locus and reviews the recent advances identified in paxilline biosynthesis.

4',6-diamidino-2-phenylindole (DAPI), netropsin, and pentamidine are minor groove binders that have terminal -C(NH2)2+ groups. The hydration changes that accompany their binding to the minor groove of the (AATT)2 sequence have been studied using the osmotic stress technique with fluorescence spectroscopy. The affinity of DAPI for the binding site decreases with the increasing osmolality of the solution, resulting in acquisition of 35+/-1 waters upon binding. A competition fluorescence assay was utilized to measure the binding constants and hydration changes of the other two ligands, using the DNA-DAPI complex as the fluorescence reporter. Upon their association to the (AATT)2 binding site, netropsin and pentamidine acquire 26+/-3 and 34+/-2 additional waters of hydration, respectively. The hydration changes are discussed in the context of the terminal functional groups of the ligands and conformational changes in the DNA.

Type 1 VWD is the mild to moderate reduction of VWF levels. This study examined the mechanisms underlying 2 common type 1 VWD mutations, the severe R1205H and more moderate Y1584C. In vitro biosynthesis was reduced for both mutations in human and mouse VWF, with the effect being more severe in R1205H. VWF knockout mice received hydrodynamic injections of mouse Vwf cDNA. Lower VWF antigen levels were demonstrated in both homozygous and heterozygous forms for both type 1 mutations from days 14-42. Recombinant protein infusions and hydrodynamicexpressed VWF propeptide to antigen ratios demonstrate that R1205H mouse VWF has an increased clearance rate, while Y1584C is normal. Recombinant AD-AMTS13 digestions of Y1584C demonstrated enhanced cleavage of both human and mouse VWF115 substrates. Hydrodynamic-expressed VWF shows a loss of high molecular weight multimers for Y1584C compared with wild-type and R1205H. At normal physiologic levels of VWF, Y1584C showed reduced thrombus formation in a ferric chloride injury model while R1205H demonstrated similar thrombogenic activity to wild-type VWF. This study has elucidated several novel mechanisms for these mutations and highlights that the type 1 VWD phenotype can be recapitulated in the VWF knockout hydrodynamic injection model. IntroductionThe large multimeric glycoprotein VWF is critical to normal hemostasis through mediating platelet-subendothelial interactions as well as binding to platelets to support their aggregation at the site of endothelial damage. The disease phenotype of type 1 VWD is a mild to moderate quantitative reduction of supposedly functionally normal VWF, with plasma VWF levels between 5% and 50% of normal. 1 This disease can be caused by a wide array of defects including defective RNA or protein synthesis, premature protein degradation before cellular release, ineffective secretion, rapid plasma clearance, or a mutation that results in a null allele. 2 R1205H, the Vicenza mutation, has a relatively severe type 1 phenotype that involves accelerated VWF clearance. Often occurring with a second VWF variation, M740I, the Vicenza mutation shows a significant reduction in VWF antigen (VWF:Ag) to ϳ 0.15 U/mL, VWF Ristocetin Cofactor Activity (VWF:RCo) ϳ 0.20 U/mL, and Factor VIII levels Ͻ 0.30 U/mL, but maintains normal platelet VWF levels and function. [3][4][5][6] Patient bleeding scores, a marker of VWD severity, range between 2-17 (n ϭ 18), with a mean of 8 (bleeding score Ն 4 is positive). 1,7-9 Accelerated clearance of the mutant protein has been demonstrated via desmopressin (DDAVP) studies 3 and human recombinant protein infusion in the VWF knockout mouse, 10 as well as through high VWFpp/ VWF:Ag ratios, with observed ratios of 10 or greater for this indirect measurement of VWF clearance from the plasma. 4 R1205H VWF also often displays an increase in high molecular weight multimers along with occasional alteration in the typical multimer triplet band pattern, [3][4][5]11 and has been attributed to the rapid clearance of the protein and thus red...

Disclaimer In an effort to expedite the publication of articles related to the COVID-19 pandemic, AJHP is posting these manuscripts online as soon as possible after acceptance. Accepted manuscripts have been peer-reviewed and copyedited, but are posted online before technical formatting and author proofing. These manuscripts are not the final version of record and will be replaced with the final article (formatted per AJHP style and proofed by the authors) at a later time. Purpose This report describes a health-system pharmacy’s response to a natural disaster while staff members simultaneously prepared for the coronavirus disease 2019 (COVID-19) pandemic. By detailing our experience, we hope to help other institutions that are current facing or could encounter similar crises. Summary In early March 2020, a tornado destroyed the health system’s warehouse for storage of most clinical supplies, including personal protective equipment and fluids. The pharmacy purchasing team collaborated with suppliers and manufacturers to recover losses and establish alternative storage areas. Days later, the pharmacy department was forced to address the impending COVID-19 pandemic. Key elements of the COVID-19 response included reducing the potential for patient and staff virus exposure; overcoming challenges in sourcing of staff, personal protective equipment, and medications; and changing care delivery practices to maintain high-quality patient care while maximizing social distancing. The pharmacy department also created distance learning opportunities for 70 pharmacy students on rotations. After an initial plan, ongoing needs include adjustment in patient care activities if significant staff losses occur, when and how to resume clinical activities, and how to best utilize the resources accumulated. Elements of practice changes implemented to reduce COVID-19 threats to patients and pharmacy personnel have proven beneficial and will be further evaluated for potential continuation. Conclusion The pharmacy department’s efforts to respond to a natural disaster and unprecedented pandemic have proven successful to this point and have illuminated several lessons, including the necessity of cohesive department communication, staff flexibility, prioritization of teamwork, and external collaboration.