Enzyme lysine 5,6 aminomutase

P-Lysine aminomutase has also been shown to catalyze the transformation of D-lysine into D-2,5-diaminohexanoate, and the enzyme has been described as o/p-lysine 5,6-aminomutase (o/P-LAM). ... 

The lysine 2,3-aminomutase reaction is reminiscent of the 1,2 rearrangement reactions catalyzed by B -dependent enzymes. The B12 dependence of several enzymic lysine 2,3-aminomutase reactions has been characterized in a variety of animals and microorganisms (222). However, the enzyme from C. subterminale SB4 is apparently not B12 requiring (220), suggesting that the enzymes catalyzing a- to j8-amino transfers are not all evolutionarily related. Given the unique cofactor requirements of the C. subterminale SB4 enzyme, i.e., SAM and Fe(ll), several studies aimed at elucidating the reaction mechanism have recently been reported. 

In contrast to these examples is lysine aminomutase, in which the only significant cluster form observed is the [4Fe-4S] form, although a [3Fe-4S] cluster can be generated upon oxidation. However, no [2Fe-2S] cluster has been reported for this enzyme. LAM is also the only member of this family in which a [4Fe-4S] " cluster has been observed. The iron-sulfur cluster in spore photoproduct lyase has not been characterized, although some evidence for [4Fe-4S] and [3Fe S] forms has been obtained. ... 

Evidence for the [4Fe S] cluster as the active form of lysine aminomutase was obtained by Frey and co-workers, who showed by a combination of EPR spectroscopy and enzyme assays that the [4Fe-4S] -LAM generated in the presence of AdoMet was catalytically active. Unlike aRNR-AE, however, LAM catalyzes a reversible reductive cleavage of AdoMet, and thus methionine production and cluster oxidation could not be monitored as evidence of turnover. It is of interest to note that in the case of LAM, the presence of AdoMet facilitates reduction to the [4Fe-4S] state very little [4Fe-4S]" cluster is produced by the reduction of LAM with dithionite in the absence of AdoMet, while the presence of AdoMet or its analogue S-adenosylhomocysteine dramatically increases the quantity of [dFe-dS] " produced. It is not clear whether the presence of AdoMet affects the redox potential of the cluster or whether some other effect, such as accessibility of the cluster by the reductant, is at work. 

Studies on three different iron-sulfur enzyme systems, which all require S-adenosyl methionine—lysine 2,3-aminomutase, pyruvate formate lyase and anaerobic ribonucleotide reductase—have led to the identification of SAM as a major source of free radicals in living cells. As in the dehydratases, these systems have a [4Fe-4S] centre chelated by only three cysteines with one accessible coordination site. The cluster is active only in the reduced... 

Recently, a new SAM dependent lysine 2,3-aminomutase was detected and characterized in Bacillus subtilis. Unlike the enzyme from C. subterminah SB4, the enzyme in B. subtilis apparently consists of four identical subunits each with a molecular mass of 54 kDa [30]. A PLP binding motif was identified in this amino-mutase that is also highly conserved in other lysine 2,3-aminomutases [31]. 

PLP radical was proved by application of [2-2H]lysine and [2-13C]lysine instead of lysine 24 to the reaction mixture [38]. In the first experiment the EPR signal was narrowed in the second it was broadened. The involvement of PLP in the reaction was proven by ESEEM spectroscopy. By incubation of the aminomutase with lysine, SAM, and [4 -2H]PLP a prominent doublet centered at the Lamour frequency for 2H was recognized [39], in accordance with the structure of an external aldi-mine. These findings establish a new role for PLP in enzyme reactions - PLP facilitates the radical isomerization. 

Besides a lysine 5,6-aminomutase, Clostridium sticklandii also has a D-ornithine 4,5-aminomutase (EC 5.4.3.5) [78, 79], D-Ornithine is generated from L-ornithine by ornithine racemase [80], The two genes encoding D-ornithine 4,5-aminomutase have been cloned, sequenced, and expressed in E. coli [81]. The enzyme is an a2/ 2-heterotetramer, consisting of 12 800 Da and 82 900 Da subunits. The protein requires Bn and pyridoxal phosphate as cofactors. Similar to the lysine 5,6-aminomutase, a conserved base-ofi)/histidine-on cobalamin binding motif is present in the 82 900 Da protein. 

So far, two types of aminomutase have been investigated in detail. Lysine 2,3-aminomutase from Clostridium subterminale SB4 is the example par excellence for the SAM-dependent type of aminomutase. Several other enzymes belonging to the same family are known. Examples are biotin synthase [82], pyruvate formate lyase [83, 84], and anaerobic ribonucleotide reductase [85]. 

Figure 10. EPR spectral evidence for SAM- or SAH-dependent reduction of the iron-sulfur clusters of lysine 2,3-aminomutase. In this figure, AdoMet refers to S-adenosylmethionine (SAM). (A) Spectrum of enzyme in the absence of dithionite and SAM (B) spectrum of enzyme in the presence of dithionite only (C) spectrum of enzyme in the presence of dithionite and SAM (D) spectrum of enzyme in the presence of dithionite and SAH. Adapted from reference 25 with permission from the American Chemical Society.

The amino mutases require pyridoxal phosphate for activity, and although little is known about them mechanistically, Freyis work on lysine 2,3-aminomutase has proved very informative (Lieder et al., 1998 Frey, 1997). Lysine 2,3-aminomutase is not a 1 2 enzyme but it functions very similarly (Section 1). The enzyme uses pyridoxal phosphate to facilitate the 1,2... 

Aminomutases. The enzymes L-p-lysine mutase (which is also D-a-lysine mutase) and D-omithine mutase catalyze the transfer of an co-amino group to an adjacent carbon atom (Table 16-1). Two proteins are needed for the reaction pyridoxal phosphate is required and is apparently directly involved in the amino group migration. In the P-lysine mutase the 6-amino group of L-P-lysine replaces the pro-S hydrogen at C-5 but with inversion at C-5 to yield (3S, 5S)-... 

The initial step in the microbiological metabolism of lysine to acetate, butyrate, and ammonia is the reversible interconversion of L-lysine and L-jS-lysine via exchange of the a-amino group and a /8-proton catalyzed by lysine 2,3-aminomutase (Scheme 49). The reaction has been observed in several species of Clostridium, Nocardia, and Streptomyces (218, 219), but the mechanistic studies have concentrated on the enzyme from Clostridium subterminale SB4, which has been purified and characterized (220). The enzyme is extremely sensitive to reversible oxygen inactivation, with reactivation achieved by anaerobic incubation with a thiol and Fe(II), and a protein-bound Fe(II) is apparently required for activity (220). The exchange reaction is also accelerated by S-... 

For many years, adenosylcobalamin was regarded as the biological source of the 5 -deoxyadenosyl radical and the most important nonoxygen source of free radicals in biocatalysis. Given the long biosynthetic pathway to adenosylcobalamin, it seemed unlikely that the unique radical chemistry of coenzyme B12 could have appeared by incremental evolution, guided solely by the survival benefits of 5 -deoxyadenosyl-5 -yl-initiated free radical chemistry in the anaerobic world. It seemed that there should have been a predecessor to adenosylcobalamin that would provide the 5 -deoxyadenosyl radical and serve as the chemical context within which adenosylcobalamin could evolve as a superior coenzyme. It seemed that a handful of SAM-dependent enzymes such as lysine 2,3-aminomutase could be the remnants of such a primordial world of radical chemistry. ° ... 

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