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Structural consequences of redesigning a protein-zinc binding site.

Ippolito JA, Christianson DW
Biochemistry 33 15241-9 (1994)
Related entries: 1cnb, 1cnc, 1cvd, 1cve, 1cvh

Cited: 19 times
EuropePMC logo PMID: 7803386

Abstract

In order to probe the structural importance of zinc ligands in the active site of human carbonic anhydrase II (CAII), we have determined the three-dimensional structures of H94C (in metal-bound form), H94C-BME (i.e., disulfide-linked with beta-mercaptoethanol), H94A, H96C, H119C, and H119D variants of CAII by X-ray crystallographic methods at resolutions of 2.2, 2.35, 2.25, 2.3, 2.2, and 2.25 A, respectively. Each variant crystallizes isomorphously with the wild-type enzyme, in which zinc is tetrahedrally coordinated by H94, H96, H119, and hydroxide ion. The structure of H94C CAII reveals the successful substitution of the naturally occurring histidine zinc ligand by a cysteine thiolate, and metal coordination by C94 is facilitated by the plastic structural response of the beta-sheet superstructure. Importantly, the resulting structure represents the catalytically active form of the enzyme reported previously [Alexander, R. S., Kiefer, L. L., Fierke, C. A., & Christianson, D. W. (1993) Biochemistry 32, 1510-1518]. Contrastingly, the structure of H96C CAII reveals that the engineered side chain does not coordinate to zinc; instead, zinc is tetrahedrally liganded by H94, H119, and two solvent molecules. Thus, the beta-sheet superstructure is not sufficiently plastic in this location to allow C96 to coordinate to the metal ion. Substitution of the thiolate or carboxylate group for wild-type histidine in H119C and H119D CAIIs reveals that tetrahedral metal coordination is maintained in each variant; however, since there is no plastic structural response of the corresponding beta-strand, a longer metal-ligand separation results.(ABSTRACT TRUNCATED AT 250 WORDS)

Reviews citing this publication (9)

  1. Neuronal CA2+/calmodulin-dependent protein kinase II: the role of structure and autoregulation in cellular function. Hudmon A, Schulman H. Annu Rev Biochem 71 473-510 (2002)
  2. Structure and mechanism of carbonic anhydrase. Lindskog S. Pharmacol Ther 74 1-20 (1997)
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  4. Catalysis by metal-activated hydroxide in zinc and manganese metalloenzymes. Christianson DW, Cox JD. Annu Rev Biochem 68 33-57 (1999)
  5. Zinc enzymes. Coleman JE. Curr Opin Chem Biol 2 222-234 (1998)
  6. Designing hydrolytic zinc metalloenzymes. Zastrow ML, Pecoraro VL. Biochemistry 53 957-978 (2014)
  7. Zinc and antibiotic resistance: metallo-beta-lactamases and their synthetic analogues. Tamilselvi A, Mugesh G. J Biol Inorg Chem 13 1039-1053 (2008)
  8. Thermodynamic, kinetic, and structural parameterization of human carbonic anhydrase interactions toward enhanced inhibitor design. Linkuvienė V, Zubrienė A, Manakova E, Petrauskas V, Baranauskienė L, Zakšauskas A, Smirnov A, Gražulis S, Ladbury JE, Matulis D. Q Rev Biophys 51 e10 (2018)
  9. Depletion and replacement of protein metal ligands. Barrick D. Curr Opin Biotechnol 6 411-418 (1995)

Articles citing this publication (10)

  1. Modulation of the phosphorylation and activity of calcium/calmodulin-dependent protein kinase II by zinc. Lengyel I, Fieuw-Makaroff S, Hall AL, Sim AT, Rostas JA, Dunkley PR. J Neurochem 75 594-605 (2000)
  2. Construction of a high affinity zinc switch in the kappa-opioid receptor. Thirstrup K, Elling CE, Hjorth SA, Schwartz TW. J Biol Chem 271 7875-7878 (1996)
  3. Selection of carbonic anhydrase variants displayed on phage. Aromatic residues in zinc binding site enhance metal affinity and equilibration kinetics. Hunt JA, Fierke CA. J Biol Chem 272 20364-20372 (1997)
  4. Molecular dynamics simulations of human carbonic anhydrase II: insight into experimental results and the role of solvation. Lu D, Voth GA. Proteins 33 119-134 (1998)
  5. ADAMDEC1 is a metzincin metalloprotease with dampened proteolytic activity. Lund J, Olsen OH, Sørensen ES, Stennicke HR, Petersen HH, Overgaard MT. J Biol Chem 288 21367-21375 (2013)
  6. Metalloprotein-inhibitor binding: human carbonic anhydrase II as a model for probing metal-ligand interactions in a metalloprotein active site. Martin DP, Hann ZS, Cohen SM. Inorg Chem 52 12207-12215 (2013)
  7. Carbon dioxide and related heterocumulenes at zinc and lithium cations: bioinspired reactions and principles. Schenk S, Notni J, Köhn U, Wermann K, Anders E. Dalton Trans 4191-4206 (2006)
  8. A combinatorial library for the binuclear metal center of bacterial phosphotriesterase. Watkins LM, Kuo JM, Chen-Goodspeed M, Raushel FM. Proteins 29 553-561 (1997)
  9. On the role of structural zinc in bis(cysteinyl) protein sequences. Meißner A, Haehnel W, Vahrenkamp H. Chemistry 3 261-267 (1997)
  10. Structural insights into the effect of active-site mutation on the catalytic mechanism of carbonic anhydrase. Kim JK, Lee C, Lim SW, Andring JT, Adhikari A, McKenna R, Kim CU. IUCrJ 7 985-994 (2020)


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