The hyaluronan-protein complexes at low ionic strength: how the hyaluronidase activity is controlled by the bovine serum albumin.
Hyaluronan (HA) hydrolysis catalysed by hyaluronidase (HAase) is strongly inhibited when performed at low HAase over HA concentration ratio and under low ionic strength conditions. The reason is the ability of long HA chains to form electrostatic and non-catalytic complexes with HAase. For a given HA concentration, low HAase concentrations lead to very low hydrolysis rates because all the HAase molecules are sequestered by HA, whilst high HAase concentrations lead to high hydrolysis rates because the excess of HAase molecules remains free and active. At pH 4, non-catalytic proteins like bovine serum albumin (BSA) are able to compete with HAase to form electrostatic complexes with HA, liberating HAase which recovers its catalytic activity. The general scheme for the BSA-dependency is thus characterised by four domains delimited by three noticeable points corresponding to constant BSA over HA concentration ratios. The existence of HA-protein complexes explains the atypical kinetic behaviour of the HA / HAase system. We also show that HAase recovers the Michaelis-Menten type behaviour when the HA molecule complexed with BSA in a constant complexion state, i.e. with the same BSA over HA ratio, is considered for substrate. When the ternary HA / HAase / BSA system is concerned, the stoichiometries of the HA-HAase and HA-BSA complexes are close to 10 protein molecules per HA molecule for a native HA of 1 MDa molar mass. Finally, we show that the behaviour of the system is similar at pH 5.25, although the efficiency of BSA is less.
Lenormand
Hélène
H
Laboratoire Polymères, Biopolymères, Surfaces, FRE 3101 CNRS, Université de Rouen, 76821 Mont-Saint-Aignan cedex, France.
Tranchepain
Frédéric
F
Deschrevel
Brigitte
B
Vincent
Jean-Claude
JC
D000818:Animals; D002417:Cattle; D006820:Hyaluronic Acid / Q000737:chemistry / Q000378:metabolism*; D006821:Hyaluronoglucosaminidase / Q000737:chemistry / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D009994:Osmolar Concentration; D012710:Serum Albumin, Bovine / Q000737:chemistry / Q000378:metabolism*; D055672:Static Electricity
D012710:Serum Albumin, Bovine; D006820:Hyaluronic Acid; D006821:Hyaluronoglucosaminidase