ORIGINAL PAPER
Characterization of the amylolytic properties of the
rumen ciliate protozoan Eudiplodinium maggii
1 |
The Kielanowski Institute of Animal Physiology and Nutrition,
Polish Academy of Sciences,
05-110 Jabłonna, Poland |
2 |
Institute of Rural Sciences University,
Aberystwyth, Llanbadarn Campus, Aberystwyth, SY23 3AL, Wales |
3 |
Rowett Research Institute,
Greenburn Road, Bucksburn, Aberdeen AB21 9SB, Scotland |
J. Anim. Feed Sci. 2007;16(4):590–606
Publication date: 2007-11-21
KEYWORDS
ABSTRACT
The rumen ciliates Eudiplodinium maggii are thought to be strongly celluloloytic. We observed,
however, that they preferentially ingested starch when the sheep were fed hay and ground barley.
The studies reported in this paper were undertaken in order to characterize the amylolytic activity
of these protozoa. The crude enzyme preparation obtained from the bacteria-free ciliates degraded
starch and dextrin at the rate of 29.5 and 19.4 μmol released glucose/mg protein/h, respectively,
while the degradation rate of maltose and isomaltose was only 0.45 and 0.14 μmol released
glucose/mg protein/h. The pH and temperature optimum of starch, dextrin, maltose and isomaltose
hydrolysis varied in the range of 4.5-7.5 and 45-55ºC relative to substrate. Pullulan was not
degraded. Four protein bands with the ability to degrade starch were identified by a zymogram
technique following the electrophoretic separation of protozoal protein. The enzymes were
α-amylase in nature, as they degraded starch mainly to maltose and maltotriose. Ion-exchange
chromatography of a crude enzyme preparation resulted in the separation of numerous fractions
which were able to degrade starch. The most amylolytic fractions were very rich in protein and also
exhibited a strong ability to digest carboxymethylcellulose. Partial sequences from two genes coding
for synthesis of α-mylase enzymes were identified in a cDNA library of Eudiplodinium maggii. The
rest of the sequences were reconstructed using GeneRacer and both complete genes were sequenced
and cloned. Gene amyl 1 consisted of 1625 bp and the amyla 2 - 1593 bp. They encoded enzymes
of 505 and 431 amino acids, respectively.
CORRESPONDING AUTHOR
G. Bełżecki
The Kielanowski Institute of Animal Physiology and Nutrition,
Polish Academy of Sciences,
05-110 Jabłonna, Poland
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