Enzyme–substrate interaction in the catalytic triad of serine proteases: increase in the pKa of Asp102

Neuvonen, Helmi

doi:10.1042/bj3220351

Cited by 3 publications

(3 citation statements)

References 38 publications

(98 reference statements)

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“…In the active site of chymotrypsin the hydroxyl group of Ser195 is hydrogen-bonded to His57, which is in turn hydrogen-bonded to Asp102 . When a peptide substrate binds to CT, a subtle change in conformation compresses the hydrogen bond between His57 and Asp102, resulting in a stronger interaction, allowing the histidine to act as an enhanced general base to abstract the proton from the Ser195 hydroxyl group.…”

Section: α-Chymotrypsin (Ct)mentioning

confidence: 99%

Overview of the Stability of α-Chymotrypsin in Different Solvent Media

2012

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Section: α-Chymotrypsin (Ct)mentioning

confidence: 99%

Overview of the Stability of α-Chymotrypsin in Different Solvent Media

2012

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“…In the D121N mutation, the change of an OH group for an NH 2 group eliminates the negative charge in the van der Waals surface of the side chain atoms. Thus, it has been suggested that the carboxyl group on the Asp would be important for the catalytic activity across different pH ranges [25] , with the protease activity of D121N being more active in a high pH environment [26] . Moreover, replacement of Asp with Glu (D121E) partially retains the colicin V secretion activity, suggesting that the carboxyl group is indeed important for the colicin V secretion.…”

Section: Discussionmentioning

confidence: 99%

Mutational Analysis of Cvab, an ABC Transporter Involved in the Secretion of Active Colicin V

Hsieh

Tai

2012

PLoS ONE

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CvaB is the central membrane transporter of the colicin V secretion system that belongs to an ATP-binding cassette superfamily. Previous data showed that the N-terminal and C-terminal domains of CvaB are essential for the function of CvaB. N-terminal domain of CvaB possesses Ca2+-dependent cysteine proteolytic activity, and two critical residues, Cys32 and His105, have been identified. In this study, we also identify Asp121 as being the third residue of the putative catalytic triad within the active site of the enzyme. The Asp121 mutants lose both their colicin V secretion activity and N-terminal proteolytic activity. The adjacent residue Pro122 also appears to play a critical role in the colicin V secretion. However, the reversal of the two residues D121P - P122D results in loss of activity. Based on molecular modeling and protein sequence alignment, several residues adjacent to the critical residues, Cys32 and His105, were also examined and characterized. Site-directed mutagenesis of Trp101, Asp102, Val108, Leu76, Gly77, and Gln26 indicate that the neighboring residues around the catalytic triad affect colicin V secretion. Several mutated CvaB proteins with defective secretion were also tested, including Asp121 and Pro122, and were found to be structurally stable. These results indicate that the residues surrounding the identified catalytic triad are functionally involved in the secretion of biologically active colicin V.

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“…3 Further, the rise of the pK a of His 57 during the catalytic process has been proposed. 11,15-18 However, as introduced recently by us, 19 there are some experimental data which indicate that the pK a of Asp 102 can also increase during the formation of the tetrahedral intermediate, and this renders possible a strong mutual interaction through hydrogen bonding between Asp 102 and His 57 .…”

Section: Introductionmentioning

confidence: 99%