Observation of Additional Calcium Ion in the Crystal Structure of the Triple Mutant K56,120,121M of Bovine Pancreatic Phospholipase A2

Rajakannan, V.; Yogavel, M.; Poi, Ming; Jeyaprakash, A. Arockia; Jeyakanthan, Jeyaraman; Velmurugan, D.; Tsai, Ming‐Daw; Sekar, K.

doi:10.1016/s0022-2836(02)01132-4

Cited by 20 publications

(34 citation statements)

References 27 publications

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“…In the present re®ned protein model three MPD molecules are found. One MPD is located in the active-site mouth and this molecule was observed in our previous studies (Sekar et al, 2003;Rajakannan et al, 2002). The remaining MPD molecules are located near the surface of the protein (Glu17/Lys108 and Ser15/Ser16); similar binding of MPD molecules was also observed in the recently reported WT structure (Steiner et al, 2001).…”

Section: Mpd Moleculessupporting

confidence: 79%

“…The r.m.s. deviations are 1.11 and 0.23 A Ê , respectively, when the present structure is superimposed (backbone atoms only) with the monoclinic form of the triple mutant (K56,120,121M; Rajakannan et al, 2002; PDB code 1gh4) and the trigonal form of the double mutant (K53,56M; Yu et al, 2000; PDB code 1c74). In the present structure, the surface-loop region (residues 60±70) is disordered, as was the case in most recombinant bovine PLA 2 structures studied so far (Sekar et al, 1997(Sekar et al, , 1998.…”

Section: Quality Of the Modelmentioning

confidence: 89%

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A redetermination of the structure of the triple mutant (K53,56,120M) of phospholipase A₂at 1.6 Å resolution using sulfur-SAS at 1.54 Å wavelength

Sekar

Rajakannan

Velmurugan

et al. 2004

Acta Crystallogr D Biol Cryst

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The crystal structure of the triple mutant K53,56,120M of bovine pancreatic phospholipase A(2) has been redetermined using sulfur single-wavelength anomalous scattering. The synchrotron data were collected at lambda = 1.54 A and the crystal diffracted to 1.6 A resolution. The program SOLVE was used to locate the heavy atoms and to estimate the initial phases and the resulting map was then subjected to RESOLVE. The output of 455 non-H atoms, including 12 S atoms, one calcium ion and one chloride ion, were then subjected to ARP/wARP followed by REFMAC. With the improved phases, the automatic model building successfully built more than 85% of the 123 residues, excluding the N- and C-terminal residues. The final crystallographic R factor is 17.7% (R(free) = 21.7%). The refined model consists of 954 non-H protein atoms, 165 water O atoms, three 2-methyl-2,4-pentanediol (MPD) molecules, one calcium ion and one chloride ion. The present work is yet another example that shows the utility of single-wavelength anomalous scattering data for solving a protein structure.

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Section: Mpd Moleculessupporting

confidence: 79%

Section: Quality Of the Modelmentioning

confidence: 89%

A redetermination of the structure of the triple mutant (K53,56,120M) of phospholipase A₂at 1.6 Å resolution using sulfur-SAS at 1.54 Å wavelength

Sekar

Rajakannan

Velmurugan

et al. 2004

Acta Crystallogr D Biol Cryst

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“…We noticed that the conserved structural water connecting the interfacial residues Ala1, Pro68, Tyr52 and Asp99 was missing. During our previous investigations of the inhibitor-free structures of other triple mutants K56,120,121M (Rajakannan et al, 2002) and K53,56,121M (Sekar et al, 2005), we observed the presence of a second calcium ion. The idea of the mutation of the residues 53, 56, 120 and 121 from lysine to methionine in the above mutants is to eliminate the anionic interface of the wild type (WT) and to enhance the zwitterionic interface (Yu et al, 2000).…”

Section: Introductionmentioning

confidence: 55%

Third calcium ion found in an inhibitor-bound phospholipase A₂

Sekar

Gayathri

Velmurugan

et al. 2006

Acta Crystallogr D Biol Cryst

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The lipolytic enzyme phospholipase A2 plays a crucial role in lipid metabolism and catalyzes hydrolysis of the fatty-acid ester bond at the sn-2 position of phospholipids. Here, the crystal structure (1.7 A resolution) of the triple mutant (K53,56,121M) of bovine pancreatic phospholipase A2 complexed with an organic molecule, p-methoxybenzoic acid (anisic acid), is reported. Residues 60-70 (the surface-loop residues) are ordered and adopt conformations which are different from those normally found in structures in which a second calcium ion is present. It is interesting to note that for the first time a third calcium ion has been identified. In addition, four Tris (2-amino-2-hydroxymethyl-1,3-propanediol) molecules were located. It is believed that one of the Tris molecules plays a role in clamping the third calcium ion and that another is involved in controlling the dynamics of the surface loop through hydrogen bonds.

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“…Recently, the crystal structure of triple mutant (K56,120,121M) reported from our laboratory (involving three of the four cationic residues) revealed the position of the secondary calcium ion for the first time in the history of bovine pancreatic PLA2. 23 In view of the above, we have undertaken the crystal structures of the free and complex forms of the triple mutant (K53,56,120M) of bovine pancreatic PLA2. The structural details and results are discussed below.…”

Section: Introductionmentioning

confidence: 99%

Crystal Structures of the Free and Anisic Acid Bound Triple Mutant of Phospholipase A2

Sekar¹,

Mala²,

Yogavel³

et al. 2003

Journal of Molecular Biology

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Observation of Additional Calcium Ion in the Crystal Structure of the Triple Mutant K56,120,121M of Bovine Pancreatic Phospholipase A2

Cited by 20 publications

References 27 publications

A redetermination of the structure of the triple mutant (K53,56,120M) of phospholipase A₂at 1.6 Å resolution using sulfur-SAS at 1.54 Å wavelength

A redetermination of the structure of the triple mutant (K53,56,120M) of phospholipase A₂at 1.6 Å resolution using sulfur-SAS at 1.54 Å wavelength

Third calcium ion found in an inhibitor-bound phospholipase A₂

Crystal Structures of the Free and Anisic Acid Bound Triple Mutant of Phospholipase A2

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Observation of Additional Calcium Ion in the Crystal Structure of the Triple Mutant K56,120,121M of Bovine Pancreatic Phospholipase A2

Cited by 20 publications

References 27 publications

A redetermination of the structure of the triple mutant (K53,56,120M) of phospholipase A2at 1.6 Å resolution using sulfur-SAS at 1.54 Å wavelength

A redetermination of the structure of the triple mutant (K53,56,120M) of phospholipase A2at 1.6 Å resolution using sulfur-SAS at 1.54 Å wavelength

Third calcium ion found in an inhibitor-bound phospholipase A2

Crystal Structures of the Free and Anisic Acid Bound Triple Mutant of Phospholipase A2

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Product

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A redetermination of the structure of the triple mutant (K53,56,120M) of phospholipase A₂at 1.6 Å resolution using sulfur-SAS at 1.54 Å wavelength

A redetermination of the structure of the triple mutant (K53,56,120M) of phospholipase A₂at 1.6 Å resolution using sulfur-SAS at 1.54 Å wavelength

Third calcium ion found in an inhibitor-bound phospholipase A₂