Biosynthesis of Peptidoglycan in Gaffkya homari. The Incorporation of Peptidoglycan into the Cell Wall and the Direction of Transpeptidation

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Abstract
Wall membrane enzyme preparations from Gaffkya homari catalyze the formation of peptidoglyean from the precursor pairs: UDP-N-acetylglucosamine + UDP-N-acetyhrluramyl-pentapeptide (UDP-MurNAc-Ala-DGlu-Lys-DAla-DAla) and also from UDP-N-acetylglucosamine + UDP-N-acetylmuramyl-tetrapeptide (UDP-MurNAc-Ala-DGlu-Lys-DAla). Part of the reaction products is soluble in 2% sodium dodecylsulfate whereas the other part is hound to pre-existing cell wall peptidoglyean. The incorporation into cell wall take place by a transpeptidation reaction in which the d-alanyl-d-alanine sequences in the pre-existing cell wall function as donors and the ∈-amino groups of the lysine residues in the newly synthesized peptidoglycan strands function as acceptors. N∈ -d-Alanyl-lysine linkages are formed. At saturating concentration of UDP-N-acetylglucosamine, the enzyme system exhibits similar apparent Km values (30-80 μM) for UDP-MurNAc-pentapeptide and UDP-MurNAc-tetrapeptide both for the formation of cell-wall-bound peptidoglycan and total (i.e. soluble + cell-wall-bound) peptidoglycan. The V values are also in the same order of magnitude (270-650 pmol × min−1× mg of protein−1). However, UDP-MurNAc-tetrapeptide was a slightly better substrate than UDP-MurNAc-pentapeptide for the formation of cell-wall-bound peptidoglycan. The synthesis of total and cell-wall-bound peptidoglycan from UDPMurNAc-pentapeptide MurNAc-pentapeptide was competitively inhibited by UDP-MurNAc-tetrapeptide and vice versa. UDP-MurNAc-tripeptide and both UDP-MurNAc-pentapeptide and UDP-MurNAc-tetrapeptide in which the ∈-amino group of the lysine residue was substituted by an acetyl group were utilized less efficiently than UDP-MurNAc-pentapeptide and UDP-MurNAc-tetrapeptide for the formation of soluble peptidoglycan; they were exceedingly poor substrates for the formation of cell-wall-bound peptidoglycan.