PMID- 12923547
OWN - NLM
STAT- MEDLINE
DCOM- 20040519
LR  - 20191210
IS  - 1087-0156 (Print)
IS  - 1087-0156 (Linking)
VI  - 21
IP  - 9
DP  - 2003 Sep
TI  - Engineering of a macromolecular scaffold to develop specific protease inhibitors.
PG  - 1063-8
AB  - The specific inhibition of serine proteases, which are crucial switches in many 
      physiologically important processes, is of value both for basic research and for 
      therapeutic applications. Ecotin, a potent macromolecular inhibitor of serine 
      proteases of the S1A family, presents an attractive scaffold to engineer specific 
      protease inhibitors because of its large inhibitor-protease interface. Using 
      synthetic shuffling in combination with a restricted tetranomial diversity, we 
      created ecotin libraries that are mutated at all 20 amino acid residues in the 
      binding interface. The efficacy of these libraries was demonstrated against the 
      serine protease plasma kallikrein (Pkal). Competitive phage display selection 
      yielded a Pkal inhibitor with an apparent dissociation equilibrium constant 
      (K(i)*) of 11 pM, whereas K(i)* values for related proteases (such as Factor Xa 
      (FXa), Factor XIa (FXIa), urokinase-type plasminogen activator (uPA), thrombin, 
      and membrane-type serine protease 1 (MT-SP1)) were four to seven orders of 
      magnitude higher. The adaptability of the scaffold was demonstrated by the 
      isolation of inhibitors to two additional serine proteases, MT-SP1/matriptase and 
      Factor XIIa.
FAU - Stoop, A Allart
AU  - Stoop AA
AD  - Department of Pharmaceutical Chemistry, University of California San Francisco, 
      600 16th Street Suite S512, San Francisco, California 94143-2280, USA.
FAU - Craik, Charles S
AU  - Craik CS
LA  - eng
GR  - CA72006/CA/NCI NIH HHS/United States
PT  - Evaluation Study
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, P.H.S.
DEP - 20030817
PL  - United States
TA  - Nat Biotechnol
JT  - Nature biotechnology
JID - 9604648
RN  - 0 (Eco protein, E coli)
RN  - 0 (Escherichia coli Proteins)
RN  - 0 (Macromolecular Substances)
RN  - 0 (Periplasmic Proteins)
RN  - 0 (Recombinant Proteins)
RN  - 0 (Serine Proteinase Inhibitors)
RN  - EC 3.4.21.- (Serine Endopeptidases)
RN  - EC 3.4.21.- (matriptase)
RN  - EC 3.4.21.34 (Plasma Kallikrein)
SB  - IM
CIN - Nat Biotechnol. 2003 Sep;21(9):1019-21. PMID: 12949566
MH  - Amino Acid Sequence
MH  - Escherichia coli Proteins/*biosynthesis/*chemistry/genetics/isolation & 
      purification
MH  - Feasibility Studies
MH  - Macromolecular Substances
MH  - Molecular Sequence Data
MH  - Molecular Weight
MH  - Mutagenesis, Site-Directed/physiology
MH  - *Periplasmic Proteins
MH  - Plasma Kallikrein/chemistry/metabolism
MH  - Protein Conformation
MH  - Protein Engineering/*methods
MH  - Recombinant Proteins/biosynthesis/chemistry/isolation & purification
MH  - Serine Endopeptidases/*chemistry/*metabolism
MH  - Serine Proteinase Inhibitors/*chemistry/genetics/isolation & 
      purification/*metabolism
EDAT- 2003/08/19 05:00
MHDA- 2004/05/20 05:00
CRDT- 2003/08/19 05:00
PHST- 2003/05/12 00:00 [received]
PHST- 2003/06/26 00:00 [accepted]
PHST- 2003/08/19 05:00 [pubmed]
PHST- 2004/05/20 05:00 [medline]
PHST- 2003/08/19 05:00 [entrez]
AID - nbt860 [pii]
AID - 10.1038/nbt860 [doi]
PST - ppublish
SO  - Nat Biotechnol. 2003 Sep;21(9):1063-8. doi: 10.1038/nbt860. Epub 2003 Aug 17.