Chain length distribution and kinetic characteristics of an enzymatically produced polymer

Mulders, K.J.M.; Beeftink, H.H.

doi:10.1515/epoly-2013-0124

Cited by 5 publications

(4 citation statements)

References 22 publications

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“…This result is in agreement with data of Mulders and Beeftink, who theoretically demonstrated the size distribution to be more prominent at higher reaction orders in HA concentration [67]. The observed increase in polydispersity over time is the effect of elongation at two separate active sites.…”

Section: Polydispersitysupporting

confidence: 93%

Structural and functional evidence for two separate oligosaccharide binding sites of Pasteurella multocida hyaluronan synthase

Kooy¹,

Beeftink²,

Eppink³

et al. 2013

AER

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Pasteurella multocida hyaluronan synthase (PmHAS) is a bi-functional glycosyltransferase, containing a β1,3-glucuronyltransferase and β1,4-N-acetylglucosaminetransferase domain. PmHAS catalyzes the elongation of hyaluronan (HA) through the sequential addition of single monosaccharides to the non-reducing end of the hyaluronan chain. Research is focused on the relation between the length of the HA oligosaccharide and the single-step elongation kinetics from HA 4 up to HA 9. It was found that the turnover number k cat increased with length to maximum values of 11 and 14 s −1 for NAc-and UA-transfer, respectively. Interestingly, the specificity constant k cat /K M increased with polymer length from HA 5 to HA 7 to a value of 44 mM −1 •s −1 , indicating an oligosaccharide binding site with increasing specificity towards a heptasaccharide at the UA domain. The value of k cat /K M remained moderately constant around 8 mM −1 •s −1 for HA 4 , HA 6 , and HA 8 , indicating a binding site with significantly lower binding specificity at the NAc domain than at the UA domain. These findings are further corroborated by a structural homology model of PmHAS, revealing two distinct sites for binding of oligosaccharides of different sizes, one in each transferase domain. Structural alignment studies between PmHAS and glycosyltransferases of the GT-A fold showed significant similarity in the binding of the UDP-sugars and the orientation of the acceptor substrate. These similarities in substrate orientation in the active site and in essential amino acid residues involved in substrate binding were utilized to localize the two HA oligosaccharide binding sites.

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Section: Polydispersitysupporting

confidence: 93%

Structural and functional evidence for two separate oligosaccharide binding sites of Pasteurella multocida hyaluronan synthase

Kooy¹,

Beeftink²,

Eppink³

et al. 2013

AER

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“…We aim at explaining macroscopic and experimentally observable quantities, such as chain length distributions (CLDs), from the underlying enzymatic mechanisms. Such CLDs can be predicted theoretically using numerical inversion of Laplace transforms [39] or kinetic equations [40][41][42]. However, besides the fact that CLDs are only one of the many quantities that can be measured in the glycogen structures we simulate, our model accounts for complex features for which an analytic treatment is no longer feasible.…”

Section: Plos Computational Biologymentioning

confidence: 99%

Stochastic modelling of a three-dimensional glycogen granule synthesis and impact of the branching enzyme

2023

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In humans, glycogen storage diseases result from metabolic inborn errors, and can lead to severe phenotypes and lethal conditions. Besides these rare diseases, glycogen is also associated to widely spread societal burdens such as diabetes. Glycogen is a branched glucose polymer synthesised and degraded by a complex set of enzymes. Over the past 50 years, the structure of glycogen has been intensively investigated. Yet, the interplay between the detailed three-dimensional glycogen structure and the related enzyme activity is only partially characterised and still to be fully understood. In this article, we develop a stochastic coarse-grained and spatially resolved model of branched polymer biosynthesis following a Gillespie algorithm. Our study largely focusses on the role of the branching enzyme, and first investigates the properties of the model with generic parameter values, before comparing it to in vivo experimental data in mice. It arises that the ratio of glycogen synthase over branching enzyme reaction rates drastically impacts the structure of the granule. We deeply investigate the mechanism of branching and parametrise it using distinct lengths. Not only do we consider various possible sets of values for these lengths, but also distinct rules to apply them. We show how combining various values for these lengths finely tunes glycogen macromolecular structure. Comparing the model with experimental data confirms that we can accurately reproduce glycogen chain length distributions in wild type mice. Additional granule properties obtained for this fit are also in good agreement with typically reported values in the experimental literature. Nonetheless, we find that the mechanism of branching must be more flexible than usually reported. Overall, our model provides a theoretical basis to quantify the effect that single enzymatic parameters, in particular of the branching enzyme, have on the chain length distribution. Our generic model and methods can be applied to any glycogen data set, and could in particular contribute to characterise the mechanisms responsible for glycogen storage disorders.

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“…Dona et al [ 46 ] reviewed in great detail in vivo and in vitro kinetic models for the digestion of starch. More recently, additional models based on Michaelis–Menten kinetics were proposed for the hydrolysis of cellulose and chitin by a processive enzyme [ 47 ] and for the polymerization of a pool of hyaluronan polymers by a nonprocessive enzyme [ 48 ]. Michaelis–Menten-based models have been successfully employed to describe metabolic pathways in aqueous environments with relatively homogeneous spatial distributions of metabolites and enzymes.…”

Section: Starch Modelingmentioning

confidence: 99%

Design starch: stochastic modeling of starch granule biogenesis

Raguin

Ebenhöh

2017

Biochemical Society Transactions

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Starch is the most widespread and abundant storage carbohydrate in plants and the main source of carbohydrate in the human diet. Owing to its remarkable properties and commercial applications, starch is still of growing interest. Its unique granular structure made of intercalated layers of amylopectin and amylose has been unraveled thanks to recent progress in microscopic imaging, but the origin of such periodicity is still under debate. Both amylose and amylopectin are made of linear chains of α-1,4-bound glucose residues, with branch points formed by α-1,6 linkages. The net difference in the distribution of chain lengths and the branching pattern of amylose (mainly linear), compared with amylopectin (racemose structure), leads to different physico-chemical properties. Amylose is an amorphous and soluble polysaccharide, whereas amylopectin is insoluble and exhibits a highly organized structure of densely packed double helices formed between neighboring linear chains. Contrarily to starch degradation that has been investigated since the early 20th century, starch production is still poorly understood. Most enzymes involved in starch growth (elongation, branching, debranching, and partial hydrolysis) are now identified. However, their specific action, their interplay (cooperative or competitive), and their kinetic properties are still largely unknown. After reviewing recent results on starch structure and starch growth and degradation enzymatic activity, we discuss recent results and current challenges for growing polysaccharides on granular surface. Finally, we highlight the importance of novel stochastic models to support the analysis of recent and complex experimental results, and to address how macroscopic properties emerge from enzymatic activity and structural rearrangements.

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Chain length distribution and kinetic characteristics of an enzymatically produced polymer

Cited by 5 publications

References 22 publications

Structural and functional evidence for two separate oligosaccharide binding sites of Pasteurella multocida hyaluronan synthase

Structural and functional evidence for two separate oligosaccharide binding sites of Pasteurella multocida hyaluronan synthase

Stochastic modelling of a three-dimensional glycogen granule synthesis and impact of the branching enzyme

Design starch: stochastic modeling of starch granule biogenesis

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