Proteases Universally Recognize Beta Strands In Their Active Sites

Tyndall, Joel D. A.; Nall, Tessa; Fairlie, David P.

doi:10.1021/cr040669e

Cited by 379 publications

(295 citation statements)

References 171 publications

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“…We found that pNA substrates with short peptide sequences (Ͻ7 residues) were very poor substrates for factor B, unlike other serine proteases that tend to recognize shorter segments of polypeptide substrates and inhibitor analogues (30,44,45). Aminomethyl courmarin substrates of Յ6 residues have been reported not to be processed effectively by C3 convertase (32).…”

Section: Discussionmentioning

confidence: 99%

Profiling the Enzymatic Properties and Inhibition of Human Complement Factor B

Abbenante

Fairlie

2007

Journal of Biological Chemistry

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Human complement factor B is the crucial catalytic component of the C3 convertase enzyme that activates the alternative pathway of complement-mediated immunity. Although a serine protease in its own right, factor B circulates in human serum as an inactive zymogen and there is a crystal structure only for the inactive state of factor B and various fragments. To provide greater insight to the catalytic function and properties of factor B, we have used short para-nitroanilide derivatives of 4-to 15-residue peptides as substrates to profile the catalytic properties of factor B. Among factors found to influence catalytic activity of factor B was an unusual dependence on pH. Non-physiological alkaline conditions strongly promoted substrate cleavage by factor B, consistent with a pH-accessible conformation of the enzyme that may be critical for catalytic function. Small N-terminal extensions to conventional hexapeptide para-nitroanilide substrates significantly increased catalytic activity of factor B, which was more selective for its cleavage site than trypsin. The new chromogenic assay enabled optimization of catalysis conditions, the profiling of different substrate sequences, and the development of the first reversible and competitive substrate-based inhibitor of factor B. The inhibitor was also shown to prevent in vitro formation of C3a from C3 by factor B, by synthetic and by natural C3 convertase of the alternative complement activation pathway, and to block formation of membrane attack complex. The availability of a reversible substrate-based inhibitor that could stabilize the active conformation of factor B, in conjunction with a pH-promoted higher processing activity, may offer a new avenue to obtain crystal structures of factor B and C3 convertase in an active conformation.

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Section: Discussionmentioning

confidence: 99%

Profiling the Enzymatic Properties and Inhibition of Human Complement Factor B

Abbenante

Fairlie

2007

Journal of Biological Chemistry

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“…Another explanation for the lack of inhibitory activity concerns the conformational rigidity of the compound. It is widely accepted that proteases generally recognize their substrates and inhibitors in an extended -strand conformation 25 . The conformation of the main chain of a peptide is described by the dihedral angles  and  (Figure 2).…”

Section: Inhibitory Activitymentioning

confidence: 99%

Dipeptide-derived nitriles containing additional electrophilic sites: Potentially irreversible inhibitors of cysteine proteases

Löser

Gütschow

2009

Journal of Enzyme Inhibition and Medicinal Chemistry

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“…A protein's susceptibility to proteolytic digestion is a functional attribute linked to its energy landscape. [8][9][10] In order to be cleaved, the substrate polypeptide chain must be extended to fit into the substrate-binding sites of a protease, 11 which make compactly folded proteins poor substrates for proteolysis. Proteolysis of compactly folded proteins requires access to high-energy cleavable states, where cleavage sites are exposed to proteases through local or global unfolding ( Fig 1A).…”

Section: Introductionmentioning

confidence: 99%

Energetics-based Protein Profiling on a Proteomic Scale: Identification of Proteins Resistant to Proteolysis

Park

Zhou

Gilmore

et al. 2007

Journal of Molecular Biology

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SUMMARYNative states of proteins are flexible, populating more than just the unique native conformation. The energetics and dynamics resulting from this conformational ensemble are inherently linked to protein function and regulation. Proteolytic susceptibility is one feature determined by this conformational energy landscape. As an attempt to investigate energetics of proteins on a proteomic scale, we challenged the E. coli proteome with extensive proteolysis and determined which proteins, if any, have optimized their energy landscape for resistance to proteolysis. To our surprise, multiple soluble proteins survived the challenge. Maltose binding protein, a survivor from thermolysin digestion, was characterized by in vitro biophysical studies to identify the physical origin of proteolytic resistance. This experimental characterization shows that kinetic stability is responsible for the unusual resistance in maltose binding protein. The biochemical functions of the identified survivors suggest that many of these proteins may have evolved extreme proteolytic resistance because of their critical roles under stressed conditions. Our results suggest that under functional selection proteins can evolve extreme proteolysis resistance by modulating their conformational energy landscapes without the need to invent new folds, and that proteins can be profiled on a proteomic scale according to their energetic properties by using proteolysis as a structural probe.

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Proteases Universally Recognize Beta Strands In Their Active Sites

Cited by 379 publications

References 171 publications

Profiling the Enzymatic Properties and Inhibition of Human Complement Factor B

Profiling the Enzymatic Properties and Inhibition of Human Complement Factor B

Dipeptide-derived nitriles containing additional electrophilic sites: Potentially irreversible inhibitors of cysteine proteases

Energetics-based Protein Profiling on a Proteomic Scale: Identification of Proteins Resistant to Proteolysis

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