Phosphonate Ester Probes for Proteolytic Antibodies

Sudhir Paul, Alfonso Tramontano, Gennady Gololobov, Yong Xin Zhou, Hiroaki Taguchi, Sangeeta Karle, Yasuhiro Nishiyama, Stephanie Planque, Saji George

Research output: Contribution to journalArticlepeer-review

64 Scopus citations


The reactivity of phosphonate ester probes with several available proteolytic antibody (Ab) fragments was characterized. Irreversible, active site-directed inhibition of the peptidase activity was evident. Stable phosphonate diester-Ab adducts were resolved by column chromatography and denaturing electrophoresis. Biotinylated phosphonate esters were applied for chemical capture of phage particles displaying Fv and light chain repertoires. Selected Ab fragments displayed enriched catalytic activity inhibitable by the selection reagent. Somewhat unexpectedly, a phosphonate monoester also formed stable adducts with the Abs. Improved catalytic activity of phage Abs selected by monoester binding was evident. Turnover values (kcat) for a selected Fv construct and a light chain against their preferred model peptide substrates were 0.5 and 0.2 min-1, respectively, and the corresponding Michaelis-Menten constants (Km) were 10 and 8 μM. The covalent reactivity of Abs with phosphonate esters suggests their ability to recapitulate the catalytic mechanism utilized by classical serine proteases.

Original languageEnglish (US)
Pages (from-to)28314-28320
Number of pages7
JournalJournal of Biological Chemistry
Issue number30
StatePublished - Jul 27 2001
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry


Dive into the research topics of 'Phosphonate Ester Probes for Proteolytic Antibodies'. Together they form a unique fingerprint.

Cite this