Temperature adaptation of proteins: engineering mesophilic-like activity and stability in a cold-adapted alpha-amylase.


Abstract

Two multiple mutants of a psychrophilic alpha-amylase were produced, bearing five mutations (each introducing additional weak interactions found in pig pancreatic alpha-amylase) with or without an extra disulfide bond specific to warm-blooded animals. Both multiple mutants display large modifications of stability and activity arising from synergic effects in comparison with single mutations. Newly introduced weak interactions and the disulfide bond confer mesophilic-like stability parameters, as shown by increases in the melting point t(m), in the calorimetric enthalpy DeltaH(cal) and in protection against heat inactivation, as well as by decreases in cooperativity and reversibility of unfolding. In addition, both kinetic and thermodynamic activation parameters of the catalyzed reaction are shifted close to the values of the porcine enzyme. This study confirms the central role of weak interactions in regulating the balance between stability and activity of an enzyme in order to adapt to the environmental temperature. Study holds ProTherm entries: 16782, 16783, 16784, 16785, 16786 Extra Details: psychrophile; cold adaptation; alpha-amylase; thermal stability; microcalorimetry

Submission Details

ID: B26v8nVV

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:48 p.m.

Version: 1

Publication Details
D'Amico S;Gerday C;Feller G,J. Mol. Biol. (2003) Temperature adaptation of proteins: engineering mesophilic-like activity and stability in a cold-adapted alpha-amylase. PMID:14499602
Additional Information

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P
 

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)


Relevant PDB Entries

Structure ID Release Date Resolution Structure Title
1HX0 2001-08-08 1.38 Structure of pig pancreatic alpha-amylase complexed with the 'truncate' acarbose molecule (pseudotrisaccharide)
1KXQ 2002-06-19 1.6 Camelid VHH Domain in Complex with Porcine Pancreatic alpha-Amylase
1KXV 2002-06-19 1.6 Camelid VHH Domains in Complex with Porcine Pancreatic alpha-Amylase
1G94 2001-12-12 1.74 CRYSTAL STRUCTURE ANALYSIS OF THE TERNARY COMPLEX BETWEEN PSYCHROPHILIC ALPHA AMYLASE FROM PSEUDOALTEROMONAS HALOPLANCTIS IN COMPLEX WITH A HEPTA-SACCHARIDE AND A TRIS MOLECULE
1G9H 2002-06-26 1.8 TERNARY COMPLEX BETWEEN PSYCHROPHILIC ALPHA-AMYLASE, COMII (PSEUDO TRI-SACCHARIDE FROM BAYER) AND TRIS (2-AMINO-2-HYDROXYMETHYL-PROPANE-1,3-DIOL)
1DHK 1997-12-24 1.85 STRUCTURE OF PORCINE PANCREATIC ALPHA-AMYLASE
1AQM 1999-03-02 1.85 ALPHA-AMYLASE FROM ALTEROMONAS HALOPLANCTIS COMPLEXED WITH TRIS
3L2M 2010-04-14 1.97 X-ray Crystallographic Analysis of Pig Pancreatic Alpha-Amylase with Alpha-cyclodextrin
1KXT 2002-06-19 2.0 Camelid VHH Domains in Complex with Porcine Pancreatic alpha-Amylase
1AQH 1999-02-16 2.0 ALPHA-AMYLASE FROM ALTEROMONAS HALOPLANCTIS
1WO2 2005-03-15 2.01 Crystal structure of the pig pancreatic alpha-amylase complexed with malto-oligosaacharides under the effect of the chloride ion
1UA3 2003-10-14 2.01 Crystal structure of the pig pancreatic a-amylase complexed with malto-oligosaccharides
1JFH 1998-12-02 2.03 STRUCTURE OF A PANCREATIC ALPHA-AMYLASE BOUND TO A SUBSTRATE ANALOGUE AT 2.03 ANGSTROM RESOLUTION
1L0P 2002-06-19 2.1 CRYSTAL STRUCTURE ANALYSIS OF THE COMPLEX BETWEEN PSYCHROPHILIC ALPHA AMYLASE FROM PSEUDOALTEROMONAS HALOPLANCTIS AND NITRATE
3L2L 2010-04-14 2.11 X-ray Crystallographic Analysis of Pig Pancreatic Alpha-Amylase with Limit Dextrin and Oligosaccharide
1PPI 1995-05-24 2.2 THE ACTIVE CENTER OF A MAMMALIAN ALPHA-AMYLASE. THE STRUCTURE OF THE COMPLEX OF A PANCREATIC ALPHA-AMYLASE WITH A CARBOHYDRATE INHIBITOR REFINED TO 2.2 ANGSTROMS RESOLUTION
1PIG 1996-12-07 2.2 PIG PANCREATIC ALPHA-AMYLASE COMPLEXED WITH THE OLIGOSACCHARIDE V-1532
1JD7 2002-09-18 2.25 CRYSTAL STRUCTURE ANALYSIS OF THE MUTANT K300R OF PSEUDOALTEROMONAS HALOPLANCTIS ALPHA-AMYLASE
1OSE 1997-04-01 2.3 Porcine pancreatic alpha-amylase complexed with acarbose
1KXH 2002-06-19 2.3 Crystal structure of the complex between an inactive mutant of psychrophilic alpha-amylase (D174N) and acarbose
1PIF 1996-12-07 2.3 PIG ALPHA-AMYLASE
1VAH 2005-04-26 2.4 Crystal structure of the pig pancreatic-amylase complexed with r-nitrophenyl-a-D-maltoside
1B0I 1999-11-17 2.4 ALPHA-AMYLASE FROM ALTEROMONAS HALOPLANCTIS
1BVN 1998-09-23 2.5 PIG PANCREATIC ALPHA-AMYLASE IN COMPLEX WITH THE PROTEINACEOUS INHIBITOR TENDAMISTAT
1JD9 2002-09-18 2.5 CRYSTAL STRUCTURE ANALYSIS OF THE MUTANT K300Q OF PSEUDOALTEROMONAS HALOPLANCTIS ALPHA-AMYLASE
4X0N 2015-11-25 2.6 Porcine pancreatic alpha-amylase in complex with helianthamide, a novel proteinaceous inhibitor

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 Alpha-amylase P29957 AMY_PSEHA
98.4 Pancreatic alpha-amylase P00690 AMYP_PIG