Homocysteine-Thiolactone - Homocysteine in Protein Structure/Function and Human Disease - page 22

Reaction 3.3 Acid and base
forms of Hcy-thiolactone
(Reprinted from [84])

+

N H 3 N H 2

- H +

O O

+ H +

S S

p K a =6.67

3.2.2 p K a value

The -amino group of Hcy-thiolactone has an unusually low pK a value of 6.67 [84],

much lower than a corresponding value for Hcy (pK a ¼

α

9.04 and 9.71) (Table 3.1 )

[192]. The lowering of pK a of an -amino group upon esterification of a carboxyl

group is a characteristic property of other

α

α -amino acid esters such as leucine

methyl ester, leucyl-tRNA (pK a ¼

7.5, [196]),
but the magnitude of the effect is less than that observed for Hcy-thiolactone.
Under physiological conditions Hcy-thiolactone exists as a neutral base form
(Reaction 3.3 ), while

7.8, [195]), or valyl-tRNA (pK a ¼

-amino group of Hcy exists in a positively charged acid
form. Because of this Hcy-thiolactone can freely diffuse through cell membranes.
In fact, in cell cultures, most of Hcy-thiolactone produced inside cells is found
extracellularly in culture media [63, 64, 67, 73, 74, 138, 190, 193, 194, 197], while
in humans and mice, most of Hcy-thiolactone produced in tissues accumulates in
the urine [93, 95].

α

3.2.3 Stability

The hydrochloric acid salt of Hcy-thiolactone is odorless and stable; it can be stored
at room temperature essentially indefinitely. In contrast, Hcy has a pungent sulfhy-
dryl odor, is less stable, and has to be stored in a freezer at 20 C. In solutions at

physiological conditions (pH 7.4, 37 C), the intramolecular thioester Hcy-
thiolactone has a half-life of ~25 h [73, 198] and is substantially more stable than
intermolecular aminoacyl-thioesters, such as methionyl-S-CoA, which hydrolyzes
with a half-life of 2.25 h [199]. The hydrolysis of Hcy-thiolactone to Hcy is much
faster in alkaline solutions. For example, in 0.1 M NaOH, Hcy-thiolactone hydro-
lysis is completed in 15 min at room temperature [75, 200].
The rate of Hcy-thiolactone hydrolysis depends on the ionization status of

its -amino group (Fig. 3.3a ) [84]. The positively charged acid form of Hcy-

thiolactone is hydrolyzed 186 times faster than the neutral base form (Table 3.2 ).
Different susceptibilities to hydrolysis of different ionic forms are characteristic of
other esters: for instance, the fully protonated form of valyl-tRNA is hydrolyzed 90
times faster than the un-protonated form [196].

α