Fluorescent properties of amino acids labeled with ortho‐aminobenzoic acid

Ito, Amando Siuiti; Turchiello, Rozane de Fátima; Hirata, Isaura Y.; Cezari, Maria Helena S.; Meldal, Morten; Juliano, Luiz

doi:10.1002/(sici)1520-6343(1998)4:6<395::aid-bspy4>3.3.co;2-p

Cited by 18 publications

(42 citation statements)

References 13 publications

(18 reference statements)

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“…It was not possible to test proline at the P2 position due to quenching of Abz fluorescence when proline is immediately adjacent to it. 9 Another series of substrates with the general sequence Abz-Leu-Xaa-# -Gln-Pro-Tyr(NO2)-Asp and 20 different amino acid residues at the P1 position were synthesized, with Leu in the P2 position. A Pro residue (instead of Gln) was placed at P2 0 to direct cleavage between Xaa and Gln, since it was believed that cleavage was unlikely to occur at P1 0 with this residue.…”

Section: Resultsmentioning

confidence: 86%

The 2.0Å Crystal Structure and Substrate Specificity of the KDEL-tailed Cysteine Endopeptidase Functioning in Programmed Cell Death of Ricinus communis Endosperm

Than

Helm

Simpson

et al. 2004

Journal of Molecular Biology

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Section: Resultsmentioning

confidence: 86%

The 2.0Å Crystal Structure and Substrate Specificity of the KDEL-tailed Cysteine Endopeptidase Functioning in Programmed Cell Death of Ricinus communis Endosperm

Than

Helm

Simpson

et al. 2004

Journal of Molecular Biology

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“…Position P1 was always occupied by Arg because TTSPs have an exclusive preference for substrates that contain this amino acid (or Lys) [2]. Amino acids at P4, to which the Abz group is linked, have no effect on the quantum yield of IQF peptides [29].…”

Section: Resultsmentioning

confidence: 99%

Probing the substrate specificities of matriptase, matriptase‐2, hepsin and DESC1 with internally quenched fluorescent peptides

Béliveau¹,

Désilets²,

Leduc³

2009

The FEBS Journal

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Type II transmembrane serine proteases (TTSPs) are a newly recognized family of S1 class proteolytic enzymes, with 20 distinct members known in mice and humans. TTSPs are divided into four subfamilies based on their modular structure [1]. The HAT ⁄ DESC sub-family is the largest and is comprised of HAT, DESC1-4 and HAT-like HATL3-5. It exhibits the simplest modular structure of the stem region, which consists of a single sea urchin sperm protein, an entero-peptidase and an agrin domain (SEA). The matriptase subfamily contains three highly homologous proteases: matriptase, matriptase-2 and matriptase-3. All matrip-tases have similar stem regions, with one SEA, two C1r ⁄ C1s, urchin embryonic growth factor, bone morphogenic protein-1 (CUB), and three (matriptase-2 and matriptase-3) or four (matriptase) low-density Type II transmembrane serine proteases are an emerging class of proteo-lytic enzymes involved in tissue homeostasis and a number of human disorders such as cancer. To better define the biochemical functions of a subset of these proteases, we compared the enzymatic properties of matriptase, matriptase-2, hepsin and DESC1 using a series of internally quenched fluorogenic peptide substrates containing o-aminobenzoyl and 3-nitro-tyro-sine. We based the sequence of the peptides on the P4 to P4¢ activation sequence of matriptase (RQAR-VVGG). Positions P4, P3, P2 and P1¢ were substituted with nonpolar (Ala, Leu), aromatic (Tyr), acid (Glu) and basic (Arg) amino acids, whereas P1 was fixed to Arg. Of the four type II trans-membrane serine proteases studied, matriptase-2 was the most promiscuous , and matriptase was the most discriminating, with a distinct specificity for Arg residues at P4, P3 and P2. DESC1 had a preference similar to that of matriptase, but with a propensity for small nonpolar amino acids (Ala) at P1¢. Hepsin shared similarities with matriptase and DESC1, but was markedly more permissive at P2. Matriptase-2 manifested broader specifici-ties, as well as substrate inhibition, for selective internally quenched fluorescent substrates. Lastly, we found that antithrombin III has robust inhibitory properties toward matriptase, matriptase-2, hepsin and DESC1, whereas plasminogen activator inhibitor-1 and a 2-antiplasmin inhibited matriptase-2, hepsin and DESC1, and to a much lesser extent, matriptase. In summary, our studies revealed that these enzymes have distinct substrate preferences. Abbreviations a 1-ACT, a 1-antichymotrypsin; AEBSF, 4-(2-aminoethyl)-benzenesulfonylfluoride hydrochloride; AMC, 7-amino-4-methylcoumarin; a 1-AP, a 1-antiplasmin;; a 1-AT, a 1-antitrypsin; AT III, antithrombin III; IQF, internally quenched fluorescent; PAI-I, plasminogen activator inhibitor I; PAR-2, protease-activated receptor-2; proMSP-1, macrophage-stimulating protein 1 precursor; PS-SCL, positional scanning-synthetic combinatorial libraries; TTSP, type II transmembrane serine protease.

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“…Two peptides were synthesized for this study: (1) ABZ‐K 2 Q 16 K 2 ‐Y (DQ 16 ) and (2) ABZ‐K 2 Q 16 K 2 ‐YNO 2 (DQ 16 A). In these peptides, ABZ denotes an n‐terminal o ‐aminobenzamide (ABZ; FRET donor), Y denotes a c‐terminal tyrosine, and YNO 2 denotes a c‐terminal 3‐nitrotyrosine (FRET acceptor) 49, 57, 58, 68. The peptides were obtained through custom synthesis (Anaspec) with purity >95% was achieved with HPLC purification and confirmed through MALDI mass spectrometry and molecular weight analysis on a Beckman XLA analytical centrifuge.…”

Section: Methodsmentioning

confidence: 99%

An accurate model of polyglutamine

et al. 2011

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Polyglutamine repeats in proteins are highly correlated with amyloid formation and neurological disease. To better understand the molecular basis of glutamine repeat diseases, structural analysis of polyglutamine peptides as soluble monomers, oligomers, and insoluble amyloid fibrils is necessary. In this study, fluorescence resonance energy transfer (FRET) experiments and molecular dynamics simulations using different theoretical models of polyglutamine were conducted. This study demonstrates that a previously proposed simple C(α)C(β) model of polyglutamine, denoted as FCO, accurately reproduced the present FRET results and the results of previously published FRET, triplet-state quenching, and fluorescence correlation studies. Other simple C(α)C(β) models with random coil and extended β-strand parameters, and all-atom models with parm96 and parm99SB force fields, did not match the FRET result well. The FCO is an intrinsically disordered model with a high-effective persistence length producing extended peptides at short lengths (Q(N) < 10). Because of an increasing number of attractive Q-Q interactions at longer lengths, the FCO model becomes increasingly more compact at lengths between Q(N) ∼ 10-16 and is as compact as many folded proteins at Q(N) > 16.

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Fluorescent properties of amino acids labeled with ortho‐aminobenzoic acid

Cited by 18 publications

References 13 publications

The 2.0Å Crystal Structure and Substrate Specificity of the KDEL-tailed Cysteine Endopeptidase Functioning in Programmed Cell Death of Ricinus communis Endosperm

The 2.0Å Crystal Structure and Substrate Specificity of the KDEL-tailed Cysteine Endopeptidase Functioning in Programmed Cell Death of Ricinus communis Endosperm

Probing the substrate specificities of matriptase, matriptase‐2, hepsin and DESC1 with internally quenched fluorescent peptides

An accurate model of polyglutamine

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