Elucidating the Role of Catalytic Amino Acid Residues in the Peptide-Mediated Silica Oligomerization Reaction Mechanism

Tropomyosins (Tpms) have been described as master regulators of actin, with Tpm3 products shown to be involved in early developmental processes, and the Tpm3 isoform Tpm3.1 controlling changes in the size of neuronal growth cones and neurite growth. Here, we used primary mouse hippocampal neurons of C57/Bl6 wild type and Bl6Tpm3flox transgenic mice to carry out morphometric analyses in response to the absence of Tpm3 products, as well as to investigate the effect of C-terminal truncation on the ability of Tpm3.1 to modulate neuronal morphogenesis. We found that the knock-out of Tpm3 leads to decreased neurite length and complexity, and that the deletion of two amino acid residues at the C-terminus of Tpm3.1 leads to more detrimental changes in neurite morphology than the deletion of six amino acid residues. We also found that Tpm3.1 that lacks the 6 C-terminal amino acid residues does not associate with stress fibres, does not segregate to the tips of neurites, and does not impact the amount of the filamentous actin pool at the axonal growth cones, as opposed to Tpm3.1, which lacks the two C-terminal amino acid residues. Our study provides further insight into the role of both Tpm3 products and the C-terminus of Tpm3.1, and it forms the basis for future studies that aim to identify the molecular mechanisms underlying Tpm3.1 targeting to different subcellular compartments.

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Role of amino acid residues at the interface of α52asparginyl-N-glycosyl chain of human choriogonadotropin

Molecular and Cellular Endocrinology ◽

10.1016/s0303-7207(99)00026-x ◽

1999 ◽

Vol 150 (1-2) ◽

pp. 47-56 ◽

Cited By ~ 1

Author(s):

Kui Shao ◽

Sathyamangalam V. Balasubramanian ◽

Om P. Bahl

Keyword(s):

Amino Acid ◽

Amino Acid Residues ◽

Human Choriogonadotropin

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The functional dyad of scorpion toxin Pi1 is not itself a prerequisite for toxin binding to the voltage-gated Kv1.2 potassium channels

Biochemical Journal ◽

10.1042/bj20030115 ◽

2004 ◽

Vol 377 (1) ◽

pp. 25-36 ◽

Cited By ~ 50

Author(s):

Stéphanie MOUHAT ◽

Amor MOSBAH ◽

Violeta VISAN ◽

Heike WULFF ◽

Muriel DELEPIERRE ◽

...

Keyword(s):

Amino Acid ◽

Scorpion Toxin ◽

Xenopus Laevis Oocytes ◽

Amino Acid Residues ◽

Voltage Gated ◽

Toxin Binding ◽

Ic50 Values

Pi1 is a 35-residue scorpion toxin cross-linked by four disulphide bridges that acts potently on both small-conductance Ca2+-activated (SK) and voltage-gated (Kv) K+ channel subtypes. Two approaches were used to investigate the relative contribution of the Pi1 functional dyad (Tyr-33 and Lys-24) to the toxin action: (i) the chemical synthesis of a [A24,A33]-Pi1 analogue, lacking the functional dyad, and (ii) the production of a Pi1 analogue that is phosphorylated on Tyr-33 (P-Pi1). According to molecular modelling, this phosphorylation is expected to selectively impact the two amino acid residues belonging to the functional dyad without altering the nature and three-dimensional positioning of other residues. P-Pi1 was directly produced by peptide synthesis to rule out any possibility of trace contamination by the unphosphorylated product. Both Pi1 analogues were compared with synthetic Pi1 for bioactivity. In vivo, [A24,A33]-Pi1 and P-Pi1 are lethal by intracerebroventricular injection in mice (LD50 values of 100 and 40 µg/mouse, respectively). In vitro, [A24,A33]-Pi1 and P-Pi1 compete with 125I-apamin for binding to SK channels of rat brain synaptosomes (IC50 values of 30 and 10 nM, respectively) and block rat voltage-gated Kv1.2 channels expressed in Xenopus laevis oocytes (IC50 values of 22 µM and 75 nM, respectively), whereas they are inactive on Kv1.1 or Kv1.3 channels at micromolar concentrations. Therefore, although both analogues are less active than Pi1 both in vivo and in vitro, the integrity of the Pi1 functional dyad does not appear to be a prerequisite for the recognition and binding of the toxin to the Kv1.2 channels, thereby highlighting the crucial role of other toxin residues with regard to Pi1 action on these channels. The computed simulations detailing the docking of Pi1 peptides on to the Kv1.2 channels support an unexpected key role of specific basic amino acid residues, which form a basic ring (Arg-5, Arg-12, Arg-28 and Lys-31 residues), in toxin binding.

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Mimicking the Function of Eggshell Matrix Proteins: The Role of Multiplets of Charged Amino Acid Residues and Self-Assembly of Peptides in Biomineralization

Angewandte Chemie International Edition ◽

10.1002/anie.200500261 ◽

2005 ◽

Vol 44 (34) ◽

pp. 5476-5479 ◽

Cited By ~ 27

Author(s):

Parayil Kumaran Ajikumar ◽

Subramanian Vivekanandan ◽

Rajamani Lakshminarayanan ◽

Seetharama D. S. Jois ◽

R. Manjunatha Kini ◽

...

Keyword(s):

Amino Acid ◽

Self Assembly ◽

Matrix Proteins ◽

Amino Acid Residues ◽

Eggshell Matrix

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Structure-activity studies of dermorphin. The role of side chains of amino acid residues on the biological activity of dermorphin

Peptides ◽

10.1016/0196-9781(84)90007-x ◽

1984 ◽

Vol 5 (4) ◽

pp. 687-689 ◽

Cited By ~ 15

Author(s):

Krzysztof Darłak ◽

Zbigniew Grzonka ◽

Pawel Krzaścik ◽

Piotr Janicki ◽

S.Witold Gumułka

Keyword(s):

Biological Activity ◽

Amino Acid ◽

Side Chains ◽

Amino Acid Residues ◽

Structure Activity

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Role of Integrase in Reverse Transcription of the Saccharomyces cerevisiae Retrotransposon Ty1

Eukaryotic Cell ◽

10.1128/ec.4.6.1057-1065.2005 ◽

2005 ◽

Vol 4 (6) ◽

pp. 1057-1065 ◽

Cited By ~ 14

Author(s):

M. Wilhelm ◽

F.-X. Wilhelm

Keyword(s):

Saccharomyces Cerevisiae ◽

Amino Acid ◽

Specific Activity ◽

Rnase H ◽

Amino Acid Residues ◽

Acidic Amino Acid ◽

Acidic Domain ◽

Basic Region

ABSTRACT Reverse transcriptase (RT) with its associated RNase H (RH) domain and integrase (IN) are key enzymes encoded by retroviruses and retrotransposons. Several studies have implied a functional role of the interaction between IN and RT during the replication of retroviral and retrotransposon genomes. In this study, IN deletion mutants were used to investigate the role of IN on the RT activity of the yeast Saccharomyces cerevisiae retrotransposon Ty1. We have identified two domains of Ty1 integrase which have effects on RT activity in vivo. The deletion of a domain spanning amino acid residues 233 to 520 of IN increases the exogenous specific activity of RT up to 20-fold, whereas the removal of a region rich in acidic amino acid residues between residues 521 and 607 decreases its activity. The last result complements our observation that an active recombinant RT protein can be obtained if a small acidic tail mimicking the acidic domain of IN is fused to the RT-RH domain. We suggest that interaction between these acidic amino acid residues of IN and a basic region of RT could be critical for the correct folding of RT and for the formation of an active conformation of the enzyme.

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Modulation of procaspase-7 self-activation by PEST amino acid residues of the N-terminal prodomain and intersubunit linker

Biochemistry and Cell Biology ◽

10.1139/bcb-2016-0220 ◽

2017 ◽

Vol 95 (6) ◽

pp. 634-643

Author(s):

Juliano Alves ◽

Miguel Garay-Malpartida ◽

João M. Occhiucci ◽

José E. Belizário

Keyword(s):

Amino Acid ◽

Large Subunit ◽

Small Subunit ◽

Cleavage Sites ◽

Amino Acid Residues ◽

Wild Type ◽

Caspase 7 ◽

Caspase Family ◽

The Impact

Procaspase-7 zymogen polypeptide is composed of a short prodomain, a large subunit (p20), and a small subunit (p10) connected to an intersubunit linker. Caspase-7 is activated by an initiator caspase-8 and -9, or by autocatalysis after specific cleavage at IQAD198↓S located at the intersubunit linker. Previously, we identified that PEST regions made of amino acid residues Pro (P), Glu (E), Asp (D), Ser (S), Thr (T), Asn (N), and Gln (Q) are conserved flanking amino acid residues in the cleavage sites within a prodomain and intersubunit linker of all caspase family members. Here we tested the impact of alanine substitution of PEST amino acid residues on procaspase-7 proteolytic self-activation directly in Escherichia coli. The p20 and p10 subunit cleavage were significantly delayed in double caspase-7 mutants in the prodomain (N18A/P26A) and intersubunit linker (S199A/P201A), compared with the wild-type caspase-7. The S199A/P201A mutants effectively inhibited the p10 small subunit cleavage. However, the mutations did not change the kinetic parameters (kcat/KM) and optimal tetrapeptide specificity (DEVD) of the purified mutant enzymes. The results suggest a role of PEST-amino acid residues in the molecular mechanism for prodomain and intersubunit cleavage and caspase-7 self-activation.

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