To the Editor:

Human hexokinase type I (EC 2.7.1.1) is the predominant glucose-phosphorylating enzyme in red blood cells. By a number of methods1 it has been proved, and is now widely accepted, that this enzyme is largely heterogeneous and present in multiple molecular forms. Hexokinase subtypes have similar kinetic properties but a different age-dependent decay and a different intracellular distribution in reticulocytes.

It is presently unknown if the multiple hexokinase subtypes reflect posttranslational modifications or different gene products. We previously showed that, at least in human placenta, the heterogeneity of hexokinase type I is caused by the presence of truncate forms arising postsynthetically.2 

In the February issue of BLOOD, Murakami and Piomelli3reported evidence for a red cell–specific hexokinase cDNA containing a unique sequence of 60 nucleotides at the beginning of the coding region. We have now identified this red cell–specific hexokinase sequence in the human genome and found that it is located 3.1 kb upstream from the somatic exon 1 (GenBank accession number AF016350). Determination of the splice-junction by direct sequencing confirmed the hypothesis that a true hexokinase isozyme may exist in humans, likely as a product of an alternative splicing event. However, human erythrocytes show a multiplicity of forms that cannot be explained only on the bases of two alternative hexokinase isoforms.4 5Thus, the origin of hexokinase multiplicity remains at least in part to be determined.

Finally, we would like to note that expression of recombinant human hexokinase type I lacking the porin-binding domain results in an enzyme with normal kinetic and regulatory properties (Bianchi et al, submitted for publication). Thus, in cases of hexokinase mutations with altered enzymatic properties, the mutation must be searched for downstream of exon 1. This exon could instead confer stability to the enzyme by favoring binding to intracellular organelles and be responsible for enzyme defects with accelerated in vivo hexokinase decay.6 

Francesca Andreoni was supported by ENEA fellowship.

1
Wilson
JE
Hexokinases.
Rev Physiol Biochem Pharmacol
126
1995
65
2
Magnani
M
Serafini
G
Bianchi
M
Casabianca
A
Stocchi
V
Human hexokinase type I microheterogeneity is due to different amino-terminal sequences.
J Biol Chem
266
1991
502
3
Murakami
K
Piomelli
S
Identification of the cDNA for human red blood cell–specific hexokinase isozyme.
Blood
89
1997
762
4
Magnani
M
Serafini
G
Stocchi
V
Hexokinase type I multiplicity in human erythrocytes.
Biochem J
254
1988
617
5
Rifken
G
Vansen
G
Kraaijenhagen
RJ
van der Vist
MJM
Vlug
AMC
Staal
GEJ
Separation and characterization of hexokinase I subtypes from human erythrocytes.
Biochem Biophys Acta
659
1981
292
6
Magnani
M
Crinelli
R
Antonelli
A
Casabianca
A
Serafini
G
The soluble but not mitochondrially bound hexokinase is a substrate for the ATP- and ubiquitin-dependent proteolytic system.
Biochem Biophys Acta
1206
1994
180
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