Cell-free preparation of carbapenems

A cell lysate comprising a group of enzymes capable of generating a compound of the Formula (I-a):

• or a salt or tautomer thereof, or a combination thereof;
• from:
• glucose and glycine or salts thereof;
• and/or
• glucose and an optionally substituted glutamate of formula (i):
• or salts thereof;
• and/or
• glucose and a proline compound of formula (iv):
• or salts thereof;
• wherein:
• the dashed line represents a single or double bond;
• each instance of R1, R2, R3, and R4 is independently selected from the group consisting of hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted heteroalkyl, optionally substituted heteroalkenyl, optionally substituted heteroalkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, and optionally substituted heteroaryl;
• R5 is selected from the group consisting of hydrogen, —OR8, —SR8, and —N(R8)2;
• each instance of R8 is independently selected from the group consisting of hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted heteroalkyl, optionally substituted heteroalkenyl, optionally substituted heteroalkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl, —C(═O)R8a, —C(═O)OR8a, —C(═O)SR8a, —C(═O)N(R8b)2, —C(═NR8b)R8a, —C(═NR8b)OR8a, —C(═NR8b)SR8a, —C(═NR8b)N(R8b)2, —C(═S)R8a, —C(═S)OR8a, —C(═S)SR8a, —C(═S)N(R8b)2, —C(═O)NR8bSO2R8a, —S(═O)R8a, —SO2R8a, —SO2N(R8a)2, —Si(R8a)3, —P(═O)(R8a)2, —P(═O)(OR8a)2, —P(═O)(R8a)(OR8a), —P(═O)(R8a)(N(R8b)2), —P(═O)(N(R8b)2)2, —P(═O)2R8a, —P(═O)2OR8a, —P(═O)2N(R8b)2, —B(R8a)2, —B(OR8a)2, and —BR8a(OR8a), wherein R8a is selected from the group consisting of optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted heteroalkyl, optionally substituted heteroalkenyl, optionally substituted heteroalkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl, if attached to an oxygen atom an oxygen protecting group, and if attached to a sulfur atom a sulfur protecting group, or two R8a groups or an R8a and R8b group are joined to form an optionally substituted heterocyclic ring;
• and each instance of R8b is independently selected from the group consisting of hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted heteroalkyl, optionally substituted heteroalkenyl, optionally substituted heteroalkynyl, optionally substituted carbocyclyl, optionally substituted heterocyclyl, optionally substituted aryl, optionally substituted heteroaryl, and a nitrogen protecting group, or two R8b are joined to form an optionally substituted heterocyclic ring;
• and
• R7a is hydrogen.