pKa Modulation of the Acid/Base Catalyst within GH32 and GH68: A Role in Substrate/Inhibitor Specificity?

Yuan, Shuguang; Roy, Katrien Le; Venken, Tom; Lammens, Willem; Ende, Wim Van den; Maeyer, Marc De

doi:10.1371/journal.pone.0037453

Cited by 20 publications

(15 citation statements)

References 53 publications

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“…The specific structure of PBAE polymer ( Fig. 1 ) used in this study was selected on the basis of initial in vitro transfection screening results in human retinal endothelial cells and RPE cells ( 28 ) and expression of a reporter gene in retina 72 hours after subretinal injection of PBAE NP carrying plasmid DNA (pDNA) ( 31 ). We now show that suprachoroidal injection of PBAE NPs results in transfection of photoreceptors and RPE throughout the entire rat eye including regions remote from the injection site.…”

Section: Discussionmentioning

confidence: 99%

Suprachoroidal gene transfer with nonviral nanoparticles

et al. 2020

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Subretinal injections of viral vectors provide great benefits but have limited cargo capacity; they induce innate and adaptive immune responses, which may cause damage and preclude repeated injections; and they pose administration risks. As a new biotechnology, suprachoroidal injections of biodegradable nanoparticles (NPs) containing a reporter plasmid induce reporter expression in rat photoreceptors and RPE throughout the entire eye and maintain expression for at least 8 months. Multiple injections markedly increase expression. Suprachoroidal injection of NPs containing a VEGF expression plasmid caused severe subretinal neovascularization progressing to subretinal fibrosis, similar to what occurs in untreated patients with neovascular age-related macular degeneration, providing a new model and proof of concept for level and duration of expression. Suprachoroidal injection of NPs containing a VEGF-binding protein expression plasmid significantly suppressed VEGF-induced vascular leakage and neovascularization demonstrating therapeutic potential. These data suggest that nonviral NP suprachoroidal gene transfer may provide a noninvasive, repeatable alternative to subretinal injection of viral vectors.

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Section: Discussionmentioning

confidence: 99%

Suprachoroidal gene transfer with nonviral nanoparticles

et al. 2020

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“…Some sugars can bind as substrates or as inhibitors in the active site of plant GH32 members (Verhaest et al, 2007) and this depends on subtle amino acid variations in the active site area. Recent pKa calculations suggest that most GH-J members show an acid–base catalyst that is not sufficiently protonated before ligand entrance, while the acid–base can be fully protonated when a substrate, but not an inhibitor, enters the catalytic pocket (Yuan et al, 2012). Moreover, the conserved arginine in the RDP motif, rather than a previously proposed Tyr in the FYASK motif, is proposed to play a key role to increase the pKa of the acid–base catalyst (Yuan et al, 2012).…”

Section: Enzymes: Structure–function Relationshipsmentioning

confidence: 99%

Multifunctional fructans and raffinose family oligosaccharides

Ende¹

2013

Front. Plant Sci.

199

116

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Fructans and raffinose family oligosaccharides (RFOs) are the two most important classes of water-soluble carbohydrates in plants. Recent progress is summarized on their metabolism (and regulation) and on their functions in plants and in food (prebiotics, antioxidants). Interest has shifted from the classic inulin-type fructans to more complex fructans. Similarly, alternative RFOs were discovered next to the classic RFOs. Considerable progress has been made in the understanding of structure–function relationships among different kinds of plant fructan metabolizing enzymes. This helps to understand their evolution from (invertase) ancestors, and the evolution and role of so-called “defective invertases.” Both fructans and RFOs can act as reserve carbohydrates, membrane stabilizers and stress tolerance mediators. Fructan metabolism can also play a role in osmoregulation (e.g., flower opening) and source–sink relationships. Here, two novel emerging roles are highlighted. First, fructans and RFOs may contribute to overall cellular reactive oxygen species (ROS) homeostasis by specific ROS scavenging processes in the vicinity of organellar membranes (e.g., vacuole, chloroplasts). Second, it is hypothesized that small fructans and RFOs act as phloem-mobile signaling compounds under stress. It is speculated that such underlying antioxidant and oligosaccharide signaling mechanisms contribute to disease prevention in plants as well as in animals and in humans.

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“…1 and 2). It was previously described that the orientation of this Trp residue is also important for Suc catalysis (Yuan et al, 2012). By altering its orientation in chicory 1-FEH IIa, a small invertase activity could be introduced (Le Roy et al, 2008).…”

Section: Introducing Invertase Activity In Nin88 By Site-directed Mutmentioning

confidence: 99%

Understanding the Role of Defective Invertases in Plants: Tobacco Nin88 Fails to Degrade Sucrose

et al. 2013

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Cell wall invertases (cwINVs), with a high affinity for the cell wall, are fundamental enzymes in the control of plant growth, development, and carbon partitioning. Most interestingly, defective cwINVs have been described in several plant species. Their highly attenuated sucrose (Suc)-hydrolyzing capacity is due to the absence of aspartate-239 (Asp-239) and tryptophan-47 (Trp-47) homologs, crucial players for stable binding in the active site and subsequent hydrolysis. However, so far, the precise roles of such defective cwINVs remain unclear. In this paper, we report on the functional characterization of tobacco (Nicotiana tabacum) Nin88, a presumed fully active cwINV playing a crucial role during pollen development. It is demonstrated here that Nin88, lacking both Asp-239 and Trp-47 homologs, has no invertase activity. This was further supported by modeling studies and site-directed mutagenesis experiments, introducing both Asp-239 and Trp-47 homologs, leading to an enzyme with a distinct Suc-hydrolyzing capacity. In vitro experiments suggest that the addition of Nin88 counteracts the unproductive and rather aspecific binding of tobacco cwINV1 to the wall, leading to higher activities in the presence of Suc and a more efficient interaction with its cell wall inhibitor. A working model is presented based on these findings, allowing speculation on the putative role of Nin88 in muro. The results presented in this work are an important first step toward unraveling the specific roles of plant defective cwINVs.

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pKa Modulation of the Acid/Base Catalyst within GH32 and GH68: A Role in Substrate/Inhibitor Specificity?

Cited by 20 publications

References 53 publications

Suprachoroidal gene transfer with nonviral nanoparticles

Suprachoroidal gene transfer with nonviral nanoparticles

Multifunctional fructans and raffinose family oligosaccharides

Understanding the Role of Defective Invertases in Plants: Tobacco Nin88 Fails to Degrade Sucrose

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