Epitope mapping and features of the epitope for monoclonal antibodies inhibiting enzymatic activity of Helicobacter pylori urease

Abstract

Two characteristic monoclonal antibodies (HpU-2 and -18) out of 26 monoclonal antibodies (HpU-1 approximately 26) produced against Helicobacter pylori (H. pylori) urease showed a strong inhibitory effect against the enzymatic activity of the urease. Epitope mapping about some monoclonal antibodies of the HpU-series inhibiting enzymatic activity was performed by using a surface plasmon resonance apparatus and by digesting H. pylori urease with trypsin, followed by mass spectroscopy. The sequences of the epitopes recognized by HpU-2 and -18 were SVELIDIGGNRRIFGFNALVDR (22 mer) and IFGFNALVDR (10 mer), respectively. The former sequence is present as a part of a loop structure at a position close to the C-terminal of the alpha-subunit of H. pylori urease, although it has been suggested that the active site of the urease resides in the beta-subunit. The above peptide (22 mer) was chemically synthesized in a linear and cyclic form, and its conjugate with BSA was immunized in rabbits. The resultant serum induced by the linear form could specifically bind to H. pylori infecting human gastric mucosa. These results suggest that the above sequence (22 mer) must be an important epitope, although it locates in the alpha-subunit but not in the beta-subunit.

Bacterial Urease and its Role in Long-Lasting Human Diseases

“…An unanticipated result was suppressing of urease activity via an allosteric effect, which might cause a distortion of the conformation of the enzyme. On the other hand, the second mechanism observed in the case of several MAb studied (which in fact possessed a weaker inhibitory effect, such as HpU-17 and -20) is assumed that MAb B binds to the vicinity of the active site, resulting in the reduction of the urease activity [140]. …”

A catalytic antibody heavy chain HpU-2 degrading its epitope peptide and H. pylori urease

“…An epitope peptide (SVELIDIGGNRRIFGFNALVD corresponding to aa 183-203 in the ␣-subunit of H. pylori urease [23]) was synthesized and used as a substrate in the cleavage reaction by the heavy chain, HpU-2-H. A 500-l solution containing purified HpU-2-H (40 g/ml) was mixed with the same volume of a solution containing 368 g/ml of the epitope peptide at 25 C in a sterilized test tube. RP-HPLC was used to monitor the time course of the cleavage of the epitope peptide.…”

Cross‐Reactivity of Polyclonal Antibodies against Canavalia ensiformis (Jack Bean) Urease and Helicobacter pylori Urease Subunit A Fragments

“…Fragment 160 – 173 showed high compatibility with the 161 – 180 fragment described in the literature . Likewise, the 190 – 202 fragment correlated well with epitopes 183 – 203 and 188 – 194 . Two additional fragments are known from the literature: 94 – 96 and 127 – 130 .…”

“…pylori recognized by antibodies against H . pylori urease include fragments: 1 – 23, 3 – 9, 18 – 32, 31 – 50, 94 – 96, 127 – 130, 161 – 180, 183 – 203, 188 – 194 …”