PMID- 19476481
OWN - NLM
STAT- MEDLINE
DCOM- 20090925
LR  - 20090803
IS  - 1742-4658 (Electronic)
IS  - 1742-464X (Linking)
VI  - 276
IP  - 13
DP  - 2009 Jul
TI  - Digestive alpha-amylases of the flour moth Ephestia kuehniella--adaptation to 
      alkaline environment and plant inhibitors.
PG  - 3531-46
LID - 10.1111/j.1742-4658.2009.07074.x [doi]
AB  - The digestive tract of lepidopteran insects is extremely alkaline. In the present 
      work, molecular adaptation of amylolytic enzymes to this environment was 
      investigated in the flour moth Ephestia kuehniella, an important stored-product 
      pest. Three digestive alpha-amylases [Ephestia kuehniella alpha-amylase 
      isoenzymes 1-3 (EkAmy1-3)] with an alkaline pH optimum were purified from larvae 
      and biochemically characterized. These isoenzymes differ significantly in their 
      sensitivity to alpha-amylase inhibitors of plant origin that are directed against 
      herbivores as antifeedants. Such functional variability renders the amylolytic 
      system less vulnerable to suppression by plant defensive molecules. Moreover, we 
      found that expression of alpha-amylases is upregulated in larvae feeding on a 
      diet enriched with an alpha-amylase inhibitor. The alpha-amylases are secreted 
      into the larval midgut by an exocytotic mechanism, as revealed by immunogold 
      microscopy. The cDNA sequence of EkAmy3 was determined, and a homology model of 
      EkAmy3 was built in order to analyze the structural features responsible for 
      adaptation to alkaline pH. First, the overall fold was found to be stabilized by 
      remodeling of ion pairs. Second, molecular simulations supported by activity 
      measurements showed that EkAmy3 does not bind a Cl(-), owing to an Arg-to-Gln 
      mutation in a conserved binding site. The Cl(-)-binding residues are in contact 
      with the catalytic residues, and this change might help to fine-tune the 
      catalytic pK(a) values to an alkaline pH optimum. We conclude that lepidopteran 
      alpha-amylases are evolutionarily adapted in terms of structure and expression 
      dynamics for effective functioning in the digestive system.
FAU - Pytelkova, Jana
AU  - Pytelkova J
AD  - Institute of Organic Chemistry and Biochemistry AS CR, v.v.i., Praha, Czech 
      Republic.
FAU - Hubert, Jan
AU  - Hubert J
FAU - Lepsik, Martin
AU  - Lepsik M
FAU - Sobotnik, Jan
AU  - Sobotnik J
FAU - Sindelka, Radek
AU  - Sindelka R
FAU - Krizkova, Iva
AU  - Krizkova I
FAU - Horn, Martin
AU  - Horn M
FAU - Mares, Michael
AU  - Mares M
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
DEP - 20090520
PL  - England
TA  - FEBS J
JT  - The FEBS journal
JID - 101229646
RN  - 0 (Insect Proteins)
RN  - 0 (Isoenzymes)
RN  - 0 (Plant Proteins)
RN  - EC 3.2.1.1 (alpha-Amylases)
SB  - IM
MH  - Amino Acid Sequence
MH  - Animals
MH  - Base Sequence
MH  - Digestive System/enzymology
MH  - *Hydrogen-Ion Concentration
MH  - Insect Proteins/chemistry/classification/genetics/*metabolism
MH  - Isoenzymes/chemistry/classification/genetics/*metabolism
MH  - Models, Molecular
MH  - Molecular Sequence Data
MH  - *Moths/anatomy & histology/embryology/enzymology
MH  - Phylogeny
MH  - Plant Proteins/genetics/*metabolism
MH  - Protein Structure, Tertiary
MH  - Sequence Alignment
MH  - Sequence Homology, Amino Acid
MH  - alpha-Amylases/chemistry/classification/genetics/*metabolism
EDAT- 2009/05/30 09:00
MHDA- 2009/09/26 06:00
CRDT- 2009/05/30 09:00
PHST- 2009/05/30 09:00 [entrez]
PHST- 2009/05/30 09:00 [pubmed]
PHST- 2009/09/26 06:00 [medline]
AID - EJB7074 [pii]
AID - 10.1111/j.1742-4658.2009.07074.x [doi]
PST - ppublish
SO  - FEBS J. 2009 Jul;276(13):3531-46. doi: 10.1111/j.1742-4658.2009.07074.x. Epub 
      2009 May 20.