A recommended procedure for the estimation of bovine testicular hyaluronidase in the presence of human serum

Gacesa, Peter; Savitsky, Marc J.; Dodgson, K. S.; Olavesen, Anthony H.

doi:10.1016/0003-2697(81)90159-7

Cited by 40 publications

(34 citation statements)

References 16 publications

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“…Despite these modifications, extrapolations to time zero were found to be impossible. Moreover, errors in the calculation of the numbers of reducing ends may result from changes in the color of the DMAB complex or its stability due to either alkaline conditions (20) or addition of ethylacetate (19), while errors may also result from the elimination of material in the centrifugation step (19) since reducing ends of HA molecules involved in complexes are eliminated from the supernatant.…”

Section: Discussionmentioning

confidence: 99%

“…Experimental solutions to this problem have been proposed. Some of those working with HA in presence of serum or proteins have introduced a step to solubilize the complex under alkaline conditions (20) or a step involving centrifugation of the sample (19) in order to avoid proteins interfering with the assay. Despite these modifications, extrapolations to time zero were found to be impossible.…”

Section: Discussionmentioning

confidence: 99%

“…Moreover, the absorbance at time zero depended on the concentration of both HA and HAase. These observations may be explained by the hypothesis in which absorbance in the Reissig et al assay is affected by HAase acting not only as an enzyme but also as a protein binding nonspecifically to HA (14,20). Gacesa et al (20) have adapted the method of Reissig et al to use the assay in the presence of serum or proteins: ethylacetate was added at the final stage of the assay and the sample was centrifuged to eliminate precipitated proteins.…”

Section: Hyaluronan (Ha)mentioning

confidence: 97%

“…The classical procedure consists in incubating a solution of HA and HAase at a constant temperature and measuring the concentration of reducing ends, via the absorbance at 585 nm, after a given incubation time (20). This procedure has been used, for example, to study HAase activity as a function of pH or ionic strength (10, 20 -24).…”

Section: Hyaluronan (Ha)mentioning

confidence: 99%

See 3 more Smart Citations

An Improved Assay for the N-Acetyl--glucosamine Reducing Ends of Polysaccharides in the Presence of Proteins

Asteriou

Deschrevel

Delpech

et al. 2001

Analytical Biochemistry

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Hyaluronan (Ha)mentioning

confidence: 97%

Section: Hyaluronan (Ha)mentioning

confidence: 99%

See 2 more Smart Citations

An Improved Assay for the N-Acetyl--glucosamine Reducing Ends of Polysaccharides in the Presence of Proteins

Asteriou

Deschrevel

Delpech

et al. 2001

Analytical Biochemistry

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“…Some other works also report that BSA can activate the HAase activity. Gacesa et al [16] report that serum proteins are able to enhance the HAase catalytic activity, and show that among the serum proteins, albumin has the greatest effect. Gold [17] observes that BSA is able to activate both human liver HAase and bovine testicular HAase at pH 4.…”

Section: Introductionmentioning

confidence: 99%

How hyaluronan–protein complexes modulate the hyaluronidase activity: The model

Vincent¹,

Lenormand²

2009

Biophysical Chemistry

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Please cite this article as: Jean-Claude Vincent, Hélène Lenormand, How hyaluronanprotein complexes modulate the hyaluronidase activity: The model, Biophysical Chemistry (2009Chemistry ( ), doi: 10.1016Chemistry ( /j.bpc.2009 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. Simulations performed by introducing experimental data produce a theoretical behaviour similar to the experimental one, including all the atypical phenomena observed : substratedependence, enzyme-dependence and protein-dependence of HAase. This shows that our assumptions are sufficient to explain the behaviour of the system and allows us to estimate unknown parameters and suggest new developments. A C C E P T E D

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Electrostatic interactions between hyaluronan and proteins at pH 4: How do they modulate hyaluronidase activity

et al. 2008

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Hyaluronan (HA) hydrolysis catalyzed by hyaluronidase (HAase) is inhibited at low HAase over HA ratio and low ionic strength, because HA forms electrostatic complexes with HAase, which is unable to catalyze hydrolysis. Bovine serum albumin (BSA) was used as a model to study the HA-protein electrostatic complexes at pH 4. At low ionic strength, there is formation of (i) neutral insoluble complexes at the phase separation and (ii) small positively-charged or large negatively-charged soluble complexes whether BSA or HA is in excess. According to the ionic strength, different types of complex are formed. Assays for HA and BSA led to the determination of the stoichiometry of these complexes. HAase was also shown to form the various types of complex with HA at low ionic strength. Finally, we showed that at 0 and 150 mmol L(-1) NaCl, BSA competes with HAase in forming complexes with HA and thus induces HAase release resulting in a large increase in the hydrolysis rate. These results, in addition to data in the literature, show that HA-protein complexes, which can exist under numerous and varied conditions of pH, ionic strength and protein over HA ratio, might control the in vivo HAase activity.

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A recommended procedure for the estimation of bovine testicular hyaluronidase in the presence of human serum

Cited by 40 publications

References 16 publications

An Improved Assay for the N-Acetyl--glucosamine Reducing Ends of Polysaccharides in the Presence of Proteins

An Improved Assay for the N-Acetyl--glucosamine Reducing Ends of Polysaccharides in the Presence of Proteins

How hyaluronan–protein complexes modulate the hyaluronidase activity: The model

Electrostatic interactions between hyaluronan and proteins at pH 4: How do they modulate hyaluronidase activity

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